From d5634e17ca812562e417b00ebe5af87cfc0962f3 Mon Sep 17 00:00:00 2001 From: Yuling1996 Date: Mon, 23 Nov 2020 16:59:16 -0600 Subject: [PATCH 01/13] spectralLibraryWriter2 --- .../SpectralLibrarySearch/LibrarySpectrum.cs | 2 + .../SpectralLibrarySearchFunction.cs | 144 ++++++++++++++++++ TaskLayer/MetaMorpheusTask.cs | 15 ++ .../SearchTask/PostSearchAnalysisTask.cs | 52 +++++++ Test/SpectralLibraryReaderTest.cs | 61 ++++++++ Test/Test.csproj | 3 + ...alLibraryWithFullsequenceFortestWriter.msp | 62 ++++++++ 7 files changed, 339 insertions(+) create mode 100644 Test/TestData/spectralLibraryWithFullsequenceFortestWriter.msp diff --git a/EngineLayer/SpectralLibrarySearch/LibrarySpectrum.cs b/EngineLayer/SpectralLibrarySearch/LibrarySpectrum.cs index 2650f5b4d..3016df168 100644 --- a/EngineLayer/SpectralLibrarySearch/LibrarySpectrum.cs +++ b/EngineLayer/SpectralLibrarySearch/LibrarySpectrum.cs @@ -17,6 +17,8 @@ public class LibrarySpectrum public int ChargeState { get; set; } public List MatchedFragmentIons { get; set; } public bool IsDecoy { get; set; } + public string ModsString { get; set; } + public string BaseSequenceWithoutMods { get; set; } public string Name { get { return Sequence + "/" + ChargeState; } diff --git a/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs b/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs index a3fd04cc3..ab94d019d 100644 --- a/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs +++ b/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs @@ -96,5 +96,149 @@ public static double CalculateNormalizedSpectralAngle(List t return normalizedSpectralAngle; } + + //1024TestDoneCompareFunction + public static double MatchedSpectraCompare(List standardSpectra, List spectraToCompare) + { + + double[] mz1 = standardSpectra.Select(b => b.Mz).ToArray(); + double intensitySum1 = standardSpectra.Select(b => b.Intensity).Sum(); + double[] intensity1 = standardSpectra.Select(b => b.Intensity / intensitySum1).ToArray(); + //Console.WriteLine(mz1.Length + " " + intensity1.Length); + Array.Sort(mz1, intensity1); + + double[] mz2 = spectraToCompare.Select(b => b.Mz).ToArray(); + double intensitySum2 = spectraToCompare.Select(b => b.Intensity).Sum(); + double[] intensity2 = spectraToCompare.Select(b => b.Intensity / intensitySum2).ToArray(); + Array.Sort(mz2, intensity2); + //Console.WriteLine(mz2.Length + " " + intensity2.Length); + + var commonNumbers = mz1.Union(mz2).ToArray(); + double min = commonNumbers.Min(); + double max = commonNumbers.Max(); + int roundMin = (int)min; + int roundMax = (int)max + 1; + //Console.WriteLine(roundMin + " " + roundMax); + + //convert spectra to vectors + List vector1 = new List(); + List vector2 = new List(); + + int i = 0; //iterate through mz1 + int k = 0; //iterate through bin + double oneMz = mz1[0]; + double oneIntensity = intensity1[0]; + //find where peaks match + while (roundMin + k * 0.5 < roundMax) + { + List x1 = new List(); + while (i < mz1.Length && roundMin + k * 0.5 <= oneMz && oneMz < roundMin + k * 0.5 + 0.5) + { + x1.Add(oneIntensity); + i++; + if (i != mz1.Length) + { + oneMz = mz1[i]; + oneIntensity = intensity1[i]; + } + } + vector1.Add(x1.Sum()); + k++; + } + + int j = 0; //iterate through mz2 + int n = 0; //iterate through bin + double twoMz = mz2[0]; + double twoIntensity = intensity2[0]; + while (roundMin + n * 0.5 < roundMax) + { + List x2 = new List(); + while (j < mz2.Length && roundMin + n * 0.5 <= twoMz && twoMz < roundMin + n * 0.5 + 0.5) + { + x2.Add(twoIntensity); + j++; + if (j != mz2.Length) + { + twoMz = mz2[j]; + twoIntensity = intensity2[j]; + } + } + vector2.Add(x2.Sum()); + n++; + } + + //numerator of dot product + double numerator = 0; + for (i = 0; i < vector1.Count; i++) + { + numerator += vector1[i] * vector2[i]; + } + + //denominator of dot product + double denominator = Math.Sqrt(vector1.Sum(x => x * x)) * Math.Sqrt(vector2.Sum(x => x * x)); + + var score = numerator / denominator; + return score; + } + + public static List AverageTwoSpectra(List spectraOne, List spectraTwo) + { + Dictionary averagedPeaksDictionary = new Dictionary(); + var averageTwoSpectraResult = new List(); + Dictionary spectraOneDictionary = new Dictionary(); + Dictionary spectraTwoDictionary = new Dictionary(); + double intensitySum1 = spectraOne.Select(b => b.Intensity).Sum(); + double intensitySum2 = spectraTwo.Select(b => b.Intensity).Sum(); + foreach (var ionOne in spectraOne) + { + var productWithChargeInOne = ionOne.NeutralTheoreticalProduct.ProductType.ToString() + ionOne.NeutralTheoreticalProduct.FragmentNumber + "^" + ionOne.Charge; + spectraOneDictionary.Add(productWithChargeInOne, ionOne); + } + foreach (var ionTwo in spectraTwo) + { + var productWithChargeInTwo = ionTwo.NeutralTheoreticalProduct.ProductType.ToString() + ionTwo.NeutralTheoreticalProduct.FragmentNumber + "^" + ionTwo.Charge; + spectraTwoDictionary.Add(productWithChargeInTwo, ionTwo); + } + var oneKeyList = spectraOneDictionary.Keys.ToList(); + var twoKeyList = spectraTwoDictionary.Keys.ToList(); + List AllproductWithCharge = new List(); + AllproductWithCharge.AddRange(oneKeyList); + foreach (var x in twoKeyList) + { + if (!AllproductWithCharge.Contains(x)) + { + AllproductWithCharge.Add(x); + } + + } + + //Tolerance tolerance = new PpmTolerance(40); + foreach (var productWithCharge in AllproductWithCharge) + { + if (spectraOneDictionary.ContainsKey(productWithCharge) && spectraTwoDictionary.ContainsKey(productWithCharge)) + { + var newMz = (spectraOneDictionary[productWithCharge].Mz + spectraTwoDictionary[productWithCharge].Mz) / 2; + var newNorIntensity = (spectraOneDictionary[productWithCharge].Intensity / intensitySum1 + spectraTwoDictionary[productWithCharge].Intensity / intensitySum2) / 2; + Product product = spectraOneDictionary[productWithCharge].NeutralTheoreticalProduct; + var newIon = new MatchedFragmentIon(ref product, newMz, newNorIntensity, spectraOneDictionary[productWithCharge].Charge); + averageTwoSpectraResult.Add(newIon); + } + else if (spectraOneDictionary.ContainsKey(productWithCharge)) + { + Product productOne = spectraOneDictionary[productWithCharge].NeutralTheoreticalProduct; + var oneIon = new MatchedFragmentIon(ref productOne, spectraOneDictionary[productWithCharge].Mz, (spectraOneDictionary[productWithCharge].Intensity / intensitySum1) / 2, spectraOneDictionary[productWithCharge].Charge); + averageTwoSpectraResult.Add(oneIon); + } + else if (spectraTwoDictionary.ContainsKey(productWithCharge)) + { + Product productTwo = spectraTwoDictionary[productWithCharge].NeutralTheoreticalProduct; + var twoIon = new MatchedFragmentIon(ref productTwo, spectraTwoDictionary[productWithCharge].Mz, (spectraTwoDictionary[productWithCharge].Intensity / intensitySum2) / 2, spectraTwoDictionary[productWithCharge].Charge); + averageTwoSpectraResult.Add(twoIon); + } + } + var testmz1 = averageTwoSpectraResult.Select(b => b.Mz).ToArray(); + var testin1 = averageTwoSpectraResult.Select(b => b.Intensity).ToArray(); + return averageTwoSpectraResult; + } } } diff --git a/TaskLayer/MetaMorpheusTask.cs b/TaskLayer/MetaMorpheusTask.cs index f4303e3c9..f1a172f1c 100644 --- a/TaskLayer/MetaMorpheusTask.cs +++ b/TaskLayer/MetaMorpheusTask.cs @@ -597,6 +597,21 @@ protected static void WritePsmsToTsv(IEnumerable psms, str } } + protected static void WriteSpectralLibrary(List spectrumLibrary, string outputFolder) + { + + string spectrumFilePath = outputFolder + "\\spectralLibrary.msp"; + using (StreamWriter output = new StreamWriter(spectrumFilePath)) + { + + foreach (var x in spectrumLibrary) + { + output.WriteLine(x.ToString()); + } + + } + } + protected void ReportProgress(ProgressEventArgs v) { OutProgressHandler?.Invoke(this, v); diff --git a/TaskLayer/SearchTask/PostSearchAnalysisTask.cs b/TaskLayer/SearchTask/PostSearchAnalysisTask.cs index f44e43a5e..506a56648 100644 --- a/TaskLayer/SearchTask/PostSearchAnalysisTask.cs +++ b/TaskLayer/SearchTask/PostSearchAnalysisTask.cs @@ -7,6 +7,7 @@ using MassSpectrometry; using MathNet.Numerics.Distributions; using Proteomics; +using Proteomics.Fragmentation; using Proteomics.ProteolyticDigestion; using System; using System.Collections.Generic; @@ -68,6 +69,7 @@ public MyTaskResults Run() HistogramAnalysis(); WritePsmResults(); + SpectralLibraryGeneration(); WriteProteinResults(); WriteQuantificationResults(); WritePrunedDatabase(); @@ -513,6 +515,56 @@ private void HistogramAnalysis() } } + //write spectral library by Yuling 11/14 + private void SpectralLibraryGeneration() + { + var FilteredPsmList = Parameters.AllPsms + .Where(p => p.FdrInfo.QValue <= 0.1 + && p.FdrInfo.QValueNotch <= CommonParameters.QValueOutputFilter).ToList(); + FilteredPsmList.RemoveAll(b => b.IsDecoy); + FilteredPsmList.RemoveAll(b => b.IsContaminant); + + //write spectral library by Yuling 11/14 + Dictionary> PsmsGroupByPeptideAndCharge = new Dictionary>(); + foreach (var x in FilteredPsmList) + { + List psmsWithsinglePeptide = Parameters.AllPsms.Where(b => b.FullSequence == x.FullSequence).OrderByDescending(p => p.Score).ToList(); + String peptideWithChargeState = x.FullSequence + "/" + x.ScanPrecursorCharge; + List psmsWithsinglePeptideAndSameCharge = psmsWithsinglePeptide.Where(b => b.ScanPrecursorCharge == x.ScanPrecursorCharge).OrderByDescending(p => p.Score).ToList(); + if (!PsmsGroupByPeptideAndCharge.ContainsKey(peptideWithChargeState)) + { + PsmsGroupByPeptideAndCharge.Add(peptideWithChargeState, psmsWithsinglePeptideAndSameCharge); + } + } + + var spectrumLibrary = new List(); + foreach (var x in PsmsGroupByPeptideAndCharge) + { + List standspctra = x.Value[0].MatchedFragmentIons; + double standPrecursurMz = x.Value[0].ScanPrecursorMonoisotopicPeakMz; + double standRt = x.Value[0].ScanRetentionTime; + int a = 0; + while (a < x.Value.Count - 1) + { + //Console.WriteLine(x.Key + " " + this.matchedSpectraCompare(standspctra, x.Value[a + 1].MatchedFragmentIons)); + var spectrumTocompare = x.Value[a + 1].MatchedFragmentIons; + + var compareScore = SpectralLibrarySearchFunction.MatchedSpectraCompare(standspctra, spectrumTocompare); + if (compareScore > 0.90) + { + standspctra = SpectralLibrarySearchFunction.AverageTwoSpectra(standspctra, spectrumTocompare); + standPrecursurMz = (standPrecursurMz + x.Value[a + 1].ScanPrecursorMonoisotopicPeakMz) / 2; + standRt = (standRt + x.Value[a + 1].ScanRetentionTime) / 2; + } + a++; + } + var standSpectrum = new LibrarySpectrum(x.Value[0].FullSequence, standPrecursurMz, x.Value[0].ScanPrecursorCharge, standspctra, standRt); + standSpectrum.BaseSequenceWithoutMods = x.Value[0].BaseSequence; + spectrumLibrary.Add(standSpectrum); + } + + WriteSpectralLibrary(spectrumLibrary, Parameters.OutputFolder); + } private void WritePsmResults() { Status("Writing PSM results...", Parameters.SearchTaskId); diff --git a/Test/SpectralLibraryReaderTest.cs b/Test/SpectralLibraryReaderTest.cs index 1c60770ca..c2ebc519c 100644 --- a/Test/SpectralLibraryReaderTest.cs +++ b/Test/SpectralLibraryReaderTest.cs @@ -148,5 +148,66 @@ public static void SpectralLibrarySearchTest() Directory.Delete(outputDir, true); } + + [Test] + public static void SpectralLibraryWriterTest() + { + var test = Path.Combine(TestContext.CurrentContext.TestDirectory, @"TestData\spectralLibraryWithFullsequenceFortestWriter.msp"); + var testLibraryWithoutDecoy = new SpectralLibrary(new List { test }); + Assert.That(testLibraryWithoutDecoy.TryGetSpectrum("AEGSDVANAVLDGADC[Common Fixed:Carbamidomethyl on C]IMLSGETAK", 3, out var spectrum)); + Assert.That(testLibraryWithoutDecoy.TryGetSpectrum("MVDENC[Common Fixed:Carbamidomethyl on C]VGFDHTVKPVSDMELETPTDK", 3, out var spectrum2)); + } + + [Test] + public static void TestSpectraCompareAndAverage() + { + var product = new Product(); + var a = new MatchedFragmentIon(ref product, 1, 2, 1); + var b = new MatchedFragmentIon(ref product, 2, 2, 1); + var c = new MatchedFragmentIon(ref product, 3, 2, 1); + var d = new MatchedFragmentIon(ref product, 4, 2, 1); + var e = new MatchedFragmentIon(ref product, 1, 4, 1); + var f = new MatchedFragmentIon(ref product, 2, 4, 1); + var g = new MatchedFragmentIon(ref product, 3, 4, 1); + var h = new MatchedFragmentIon(ref product, 5, 4, 1); + + var compare1 = SpectralLibrarySearchFunction.MatchedSpectraCompare(new List { a, b, c }, new List { f, g, h }); + var compare2 = SpectralLibrarySearchFunction.MatchedSpectraCompare(new List { a, b, c }, new List { a, b, c }); + var compare3 = SpectralLibrarySearchFunction.MatchedSpectraCompare(new List { a, b, h }, new List { f, e, g }); + Assert.That(Math.Abs(compare1 - 0.6667) < 0.001); + Assert.That(Math.Abs(compare2 - 1) < 0.001); + Assert.That(Math.Abs(compare3 - 0.4714) < 0.001); + + //test averageTwoSpectraFunction by Yuling + Product b1 = new Product(ProductType.b, FragmentationTerminus.Both, 0, 1, 0, 0); + Product b2 = new Product(ProductType.b, FragmentationTerminus.Both, 0, 2, 0, 0); + Product b3 = new Product(ProductType.b, FragmentationTerminus.Both, 0, 3, 0, 0); + Product b4 = new Product(ProductType.b, FragmentationTerminus.Both, 0, 4, 0, 0); + Product b5 = new Product(ProductType.b, FragmentationTerminus.Both, 0, 5, 0, 0); + Product y1 = new Product(ProductType.y, FragmentationTerminus.Both, 0, 1, 0, 0); + Product y2 = new Product(ProductType.y, FragmentationTerminus.Both, 0, 2, 0, 0); + Product y3 = new Product(ProductType.y, FragmentationTerminus.Both, 0, 3, 0, 0); + Product y4 = new Product(ProductType.y, FragmentationTerminus.Both, 0, 4, 0, 0); + Product y5 = new Product(ProductType.y, FragmentationTerminus.Both, 0, 5, 0, 0); + + var o = new MatchedFragmentIon(ref b1, 1, 3, 1); + var p = new MatchedFragmentIon(ref b2, 2, 2, 1); + var q = new MatchedFragmentIon(ref b3, 3, 1, 1); + var r = new MatchedFragmentIon(ref b1, 4, 2, 2); + var s = new MatchedFragmentIon(ref b2, 2, 4, 1); + var t = new MatchedFragmentIon(ref b3, 2, 4, 2); + + // all same charge + var ave1 = SpectralLibrarySearchFunction.AverageTwoSpectra(new List { o, p, q }, new List { o, p, q }); + Assert.AreEqual(ave1.Count, 3); + Assert.AreEqual(ave1[0], new MatchedFragmentIon(ref b1, 1, 0.5, 1)); + + // with different charges + var ave2 = SpectralLibrarySearchFunction.AverageTwoSpectra(new List { o, p, q }, new List { r, s, t }); + Assert.AreEqual(ave2.Count, 5); + Assert.AreEqual(ave2[0].Intensity, 0.25); + Assert.That(Math.Abs(ave2[1].Intensity - 0.3667) < 0.01); + + } } } diff --git a/Test/Test.csproj b/Test/Test.csproj index 64ffad5b5..102906f82 100644 --- a/Test/Test.csproj +++ b/Test/Test.csproj @@ -205,6 +205,9 @@ Always + + Always + Always diff --git a/Test/TestData/spectralLibraryWithFullsequenceFortestWriter.msp b/Test/TestData/spectralLibraryWithFullsequenceFortestWriter.msp new file mode 100644 index 000000000..a6b60415d --- /dev/null +++ b/Test/TestData/spectralLibraryWithFullsequenceFortestWriter.msp @@ -0,0 +1,62 @@ +Name: AEGSDVANAVLDGADC[Common Fixed:Carbamidomethyl on C]IMLSGETAK/3 +MW: 832.0530474967455 +Comment: Parent=832.0530474967455 RT=179.79274259073895 +Num peaks: 30 +147.11208963394165 0.06901563868513455 "y1^1/0ppm" +201.0862011909485 0.00013797485462015792 "b2^1/0ppm" +218.14799094200134 0.00044056506519407954 "y2^1/0ppm" +258.10840702056885 2.875761111635475E-05 "b3^1/0ppm" +319.1983485817909 0.0026598563194152624 "y3^1/0ppm" +345.1388473510742 3.44645820813353E-06 "b4^1/0ppm" +448.23669028282166 0.0001427632781793673 "y4^1/0ppm" +460.1653690338135 9.949696016589893E-05 "b5^1/0ppm" +505.25952408229386 0.2309473308837164 "y5^1/0ppm" +559.2320953831266 0.0006791786549231851 "b6^1/0ppm" +592.2889990419209 0.569813212278294 "y6^1/0ppm" +630.2676391601562 0.0014102099194122576 "b7^1/0ppm" +705.3722317279978 0.5001700835173554 "y7^1/0ppm" +744.3114669414258 0.0002125164655079635 "b8^1/0ppm" +836.4131565672677 1 "y8^1/0ppm" +815.3434219650169 0.03768849419691366 "b9^1/0ppm" +949.504334016519 0.02640559401410081 "y9^1/0ppm" +914.4125983714149 0.141788939426318 "b10^1/0ppm" +1109.5309445698574 0.001991982494868762 "y10^1/0ppm" +1027.5084172848406 0.0011620930946783098 "b11^1/0ppm" +1224.5530804202501 0.00014674952510367384 "y11^1/0ppm" +1142.5369750519822 0.10394671678596142 "b12^1/0ppm" +1199.5529479980469 2.3227938504710007E-06 "b13^1/0ppm" +1352.6060485839844 1.9338787552222737E-06 "y13^1/0ppm" +635.7938376519564 0.00011625370225527281 "b14^2/0ppm" +1467.6546020507812 2.229388701866764E-06 "y14^1/0ppm" +1385.626220703125 9.037963503814786E-07 "b15^1/0ppm" +790.865859741715 1.0571022077310286E-05 "y15^2/0ppm" +1580.75341796875 4.770517990550197E-06 "y15^1/0ppm" +1270.5919189453125 5.642230420303506E-06 "b14^1/0ppm" +Name: MVDENC[Common Fixed:Carbamidomethyl on C]VGFDHTVKPVSDMELETPTDK/3 +MW: 1031.8014564088521 +Comment: Parent=1031.8014564088521 RT=115.556645 +Num peaks: 24 +147.11217498779297 0.169782191451439 "y1^1/0ppm" +231.1152572631836 0.7166405723415292 "b2^1/0ppm" +262.1389617919922 0.15465406562007505 "y2^1/0ppm" +346.1414031982422 0.5655131364819993 "b3^1/0ppm" +475.1841735839844 0.28176401902446185 "b4^1/0ppm" +460.23877039989037 1 "y4^1/0ppm" +589.2265014648438 0.3166002815219039 "b5^1/0ppm" +561.284834861942 0.5877647394176805 "y5^1/0ppm" +749.2506103515625 0.29131651888173665 "b6^1/0ppm" +690.3282165527344 0.2144213590855243 "y6^1/0ppm" +848.3348388671875 0.11273975514773636 "b7^1/0ppm" +803.4120483398438 0.21515612454141553 "y7^1/0ppm" +932.45849609375 0.1349192914905513 "y8^1/0ppm" +1063.4822387695312 0.1904409822149939 "y9^1/0ppm" +1461.6809692382812 0.2487322979467983 "y13^1/0ppm" +816.8644009441363 0.07191830160284918 "b14^2/0ppm" +795.3865251471564 0.22894024178847877 "y14^2/0ppm" +1015.951004544102 0.09562912185995111 "b18^2/0ppm" +1123.5296408369852 0.92028111665632 "y20^2/0ppm" +873.7331801994648 0.09559716822610352 "y23^3/0ppm" +916.7477753815713 0.11232281047234934 "y24^3/0ppm" +955.0930534844096 0.9736671274995223 "y25^3/0ppm" +988.1123439504893 0.32119688665543367 "y26^3/0ppm" +363.1920166015625 0.05089008577644779 "y3^1/0ppm" \ No newline at end of file From 3dbb4826b1f3b0f02e68be7e63f312eb2598ff62 Mon Sep 17 00:00:00 2001 From: trishorts Date: Tue, 24 Nov 2020 09:00:13 -0600 Subject: [PATCH 02/13] add spectralLibrary.msp to expected file output --- Test/MyTaskTest.cs | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/Test/MyTaskTest.cs b/Test/MyTaskTest.cs index 5f477e46c..ac70ed0a9 100644 --- a/Test/MyTaskTest.cs +++ b/Test/MyTaskTest.cs @@ -458,7 +458,7 @@ public static void TestFileOutput() // test single file output. With only 1 file, all results including mzid get written to the parent directory. HashSet expectedFiles = new HashSet { "AllPeptides.psmtsv", "AllProteinGroups.tsv", "AllPSMs.psmtsv", "AllPSMs_FormattedForPercolator.tab", "AllQuantifiedPeaks.tsv", - "AllQuantifiedPeptides.tsv", "prose.txt", "results.txt", "sliced-raw.mzID" }; + "AllQuantifiedPeptides.tsv", "prose.txt", "results.txt", "sliced-raw.mzID", "spectralLibrary.msp" }; HashSet files = new HashSet(Directory.GetFiles(Path.Combine(thisTaskOutputFolder, "SingleMassSpectraFileOutput")).Select(v => Path.GetFileName(v))); @@ -472,7 +472,7 @@ public static void TestFileOutput() // test multi file output. With multiple files, .mzid results are inside a folder. expectedFiles = new HashSet { "AllPeptides.psmtsv", "AllProteinGroups.tsv", "AllPSMs.psmtsv", "AllPSMs_FormattedForPercolator.tab", "AllQuantifiedPeaks.tsv", - "AllQuantifiedPeptides.tsv", "prose.txt", "results.txt"}; + "AllQuantifiedPeptides.tsv", "prose.txt", "results.txt", "spectralLibrary.msp"}; files = new HashSet(Directory.GetFiles(Path.Combine(thisTaskOutputFolder, "MultipleMassSpectraFileOutput")).Select(v => Path.GetFileName(v))); missingFiles = expectedFiles.Except(files); extraFiles = files.Except(expectedFiles); From ea8f0cfc361f42a46b31e5abc7c88366dd7ee7e5 Mon Sep 17 00:00:00 2001 From: trishorts Date: Tue, 24 Nov 2020 09:36:20 -0600 Subject: [PATCH 03/13] fix unit test my adding a fragment ion --- Test/gptmdPrunedBdTests.cs | 8 +++++--- 1 file changed, 5 insertions(+), 3 deletions(-) diff --git a/Test/gptmdPrunedBdTests.cs b/Test/gptmdPrunedBdTests.cs index c368c08f8..f4054e53a 100644 --- a/Test/gptmdPrunedBdTests.cs +++ b/Test/gptmdPrunedBdTests.cs @@ -358,8 +358,9 @@ public static void TestProteinPrunedWithModSelection() PostSearchAnalysisParameters testPostTaskParameters = new PostSearchAnalysisParameters(); CommonParameters commonParam = new CommonParameters(useDeltaScore: false); double[,] noiseData = new double[10000, 10000]; - noiseData[0,0] = 1.0; - List matchedFragmentIons = new List() { }; + noiseData[0,0] = 1.0; + var product = new Proteomics.Fragmentation.Product(); + List matchedFragmentIons = new List() { new Proteomics.Fragmentation.MatchedFragmentIon(ref product,1,2,1) }; MzSpectrum spectrum = new MzSpectrum(noiseData); MsDataScan scan = new MsDataScan(spectrum , 1, 1, true, Polarity.Unknown, 2, new MzLibUtil.MzRange(10, 1000), "", MZAnalyzerType.Orbitrap, 10000, null, noiseData, ""); testPostTaskParameters.ProteinList = new List() { protein1, protein2 }; @@ -464,7 +465,8 @@ public static void TestProteinPrunedWithModSelectionAndVariants() CommonParameters commonParam = new CommonParameters(useDeltaScore: false); double[,] noiseData = new double[10000, 10000]; noiseData[0, 0] = 1.0; - List matchedFragmentIons = new List() { }; + var product = new Proteomics.Fragmentation.Product(); + List matchedFragmentIons = new List() { new Proteomics.Fragmentation.MatchedFragmentIon(ref product, 1, 2, 1) }; MzSpectrum spectrum = new MzSpectrum(noiseData); MsDataScan scan = new MsDataScan(spectrum, 1, 1, true, Polarity.Unknown, 2, new MzLibUtil.MzRange(10, 1000), "", MZAnalyzerType.Orbitrap, 10000, null, noiseData, ""); testPostTaskParameters.ProteinList = proteinList; From 410f6c5d89102d1a167ef69346782f2fb06cc340 Mon Sep 17 00:00:00 2001 From: trishorts Date: Tue, 24 Nov 2020 09:39:08 -0600 Subject: [PATCH 04/13] add spectralLibrary.msp to expect files in variant search --- Test/VariantSearchTests.cs | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/Test/VariantSearchTests.cs b/Test/VariantSearchTests.cs index 16c0d619d..a652846a9 100644 --- a/Test/VariantSearchTests.cs +++ b/Test/VariantSearchTests.cs @@ -192,7 +192,7 @@ public void VariantSpecificOutputFiles() // no variant results files should be generated HashSet expectedFiles = new HashSet { - "AllPeptides.psmtsv", "AllPSMs.psmtsv", "AllPSMs_FormattedForPercolator.tab", "prose.txt", "results.txt" }; + "AllPeptides.psmtsv", "AllPSMs.psmtsv", "AllPSMs_FormattedForPercolator.tab", "prose.txt", "results.txt", "spectralLibrary.msp" }; HashSet files1 = new HashSet(Directory.GetFiles(Path.Combine(thisTaskOutputFolder, "NoVariantOutput")).Select(v => Path.GetFileName(v))); @@ -206,7 +206,7 @@ public void VariantSpecificOutputFiles() HashSet files2 = new HashSet(Directory.GetFiles(Path.Combine(thisTaskOutputFolder, "VariantOutput_frameshift")).Select(v => Path.GetFileName(v))); // variant files should be generates expectedFiles = new HashSet { - "AllPeptides.psmtsv", "AllPSMs.psmtsv", "AllPSMs_FormattedForPercolator.tab", "AllProteinGroups.tsv", "AllQuantifiedPeaks.tsv", "AllQuantifiedPeptides.tsv", "prose.txt", "results.txt", "TestVariantPep.mzID", "VariantPeptides.psmtsv", "VariantAnalysisResultSummary.txt", "VariantPSMs.psmtsv" }; + "AllPeptides.psmtsv", "AllPSMs.psmtsv", "AllPSMs_FormattedForPercolator.tab", "AllProteinGroups.tsv", "AllQuantifiedPeaks.tsv", "AllQuantifiedPeptides.tsv", "prose.txt", "results.txt", "TestVariantPep.mzID", "VariantPeptides.psmtsv", "VariantAnalysisResultSummary.txt", "VariantPSMs.psmtsv", "spectralLibrary.msp" }; // these 2 lines are for debug purposes, so you can see which files you're missing (if any) missingFiles = expectedFiles.Except(files2).ToList(); From af1d5afba612f6754a6c37a4ade35d5401f75172 Mon Sep 17 00:00:00 2001 From: Yuling1996 Date: Sun, 29 Nov 2020 18:16:08 -0600 Subject: [PATCH 05/13] 1129SPECTRALlIBRARYSEARCH --- .../ClassicSearch/ClassicSearchEngine.cs | 8 +- .../SpectralLibrarySearch/SpectralLibrary.cs | 10 +- .../SpectralLibrarySearchFunction.cs | 90 +- latest1129pepMTlibrary.csv | 52015 ++++++++++++++++ latest1129pepMTlibrary.xlsx | Bin 0 -> 501273 bytes 5 files changed, 52108 insertions(+), 15 deletions(-) create mode 100644 latest1129pepMTlibrary.csv create mode 100644 latest1129pepMTlibrary.xlsx diff --git a/EngineLayer/ClassicSearch/ClassicSearchEngine.cs b/EngineLayer/ClassicSearch/ClassicSearchEngine.cs index 00ec7e2ea..3200ceee4 100644 --- a/EngineLayer/ClassicSearch/ClassicSearchEngine.cs +++ b/EngineLayer/ClassicSearch/ClassicSearchEngine.cs @@ -79,10 +79,10 @@ protected override MetaMorpheusEngineResults RunSpecific() foreach (ScanWithIndexAndNotchInfo scan in GetAcceptableScans(peptide.MonoisotopicMass, SearchMode)) { - if (SpectralLibrary != null && !SpectralLibrary.ContainsSpectrum(peptide.FullSequence, scan.TheScan.PrecursorCharge)) - { - continue; - } + //if (SpectralLibrary != null && !SpectralLibrary.ContainsSpectrum(peptide.FullSequence, scan.TheScan.PrecursorCharge)) + //{ + //continue; + //} List matchedIons = MatchFragmentIons(scan.TheScan, peptideTheorProducts, CommonParameters); diff --git a/EngineLayer/SpectralLibrarySearch/SpectralLibrary.cs b/EngineLayer/SpectralLibrarySearch/SpectralLibrary.cs index 725fcf024..8f8b731b0 100644 --- a/EngineLayer/SpectralLibrarySearch/SpectralLibrary.cs +++ b/EngineLayer/SpectralLibrarySearch/SpectralLibrary.cs @@ -185,7 +185,15 @@ private LibrarySpectrum ReadLibrarySpectrum(StreamReader reader, bool onlyReadHe sequence = split[0].Replace("Name:", string.Empty).Trim(); // get charge - z = int.Parse(split[1].Trim()); + try + { + z = int.Parse(split[1].Trim()); + } + catch + { + var chargeLine = split[1].ToCharArray(); + z = int.Parse(chargeLine[0].ToString()); + } } else if (line.StartsWith("MW", StringComparison.InvariantCultureIgnoreCase)) { diff --git a/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs b/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs index ab94d019d..9174c5954 100644 --- a/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs +++ b/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs @@ -2,6 +2,7 @@ using MassSpectrometry; using MzLibUtil; using Proteomics.Fragmentation; +using Proteomics.ProteolyticDigestion; using System; using System.Collections.Generic; using System.IO; @@ -22,14 +23,20 @@ public static void CalculateSpectralAngles(SpectralLibrary spectralLibrary, Pept //TODO: spectral angle could be used to disambiguate PSMs. right now for ambiguous PSMs, the spectral angle for only one peptide option is saved foreach (var peptide in psm.PeptidesToMatchingFragments) { - if (spectralLibrary == null || !spectralLibrary.TryGetSpectrum(peptide.Key.FullSequence, scan.PrecursorCharge, out var librarySpectrum)) + if (spectralLibrary != null && peptide.Key.Protein.IsDecoy && !spectralLibrary.ContainsSpectrum(peptide.Key.FullSequence, scan.PrecursorCharge)) { - continue; + double spectralAngle = CalculateDecoyNormalizedSpectralAngle(peptide.Key, scan, commonParameters); + psm.SpectralAngle = spectralAngle; + } + else if (spectralLibrary != null && spectralLibrary.TryGetSpectrum(peptide.Key.FullSequence, scan.PrecursorCharge, out var librarySpectrum)) + { + double spectralAngle = CalculateNormalizedSpectralAngle(librarySpectrum.MatchedFragmentIons, scan.TheScan, commonParameters); + psm.SpectralAngle = spectralAngle; + } + else + { + psm.SpectralAngle = -1; } - - double spectralAngle = CalculateNormalizedSpectralAngle(librarySpectrum.MatchedFragmentIons, scan.TheScan, commonParameters); - - psm.SpectralAngle = spectralAngle; } } } @@ -103,13 +110,13 @@ public static double MatchedSpectraCompare(List standardSpec double[] mz1 = standardSpectra.Select(b => b.Mz).ToArray(); double intensitySum1 = standardSpectra.Select(b => b.Intensity).Sum(); - double[] intensity1 = standardSpectra.Select(b => b.Intensity / intensitySum1).ToArray(); + double[] intensity1 = standardSpectra.Select(b => Math.Sqrt(b.Intensity / intensitySum1)).ToArray(); //Console.WriteLine(mz1.Length + " " + intensity1.Length); Array.Sort(mz1, intensity1); double[] mz2 = spectraToCompare.Select(b => b.Mz).ToArray(); double intensitySum2 = spectraToCompare.Select(b => b.Intensity).Sum(); - double[] intensity2 = spectraToCompare.Select(b => b.Intensity / intensitySum2).ToArray(); + double[] intensity2 = spectraToCompare.Select(b => Math.Sqrt(b.Intensity / intensitySum2)).ToArray(); Array.Sort(mz2, intensity2); //Console.WriteLine(mz2.Length + " " + intensity2.Length); @@ -226,13 +233,13 @@ public static List AverageTwoSpectra(List AverageTwoSpectra(List b.Intensity).ToArray(); return averageTwoSpectraResult; } + + public static double CalculateDecoyNormalizedSpectralAngle(PeptideWithSetModifications peptide, Ms2ScanWithSpecificMass scan, CommonParameters commonParameters) + + { + var theoreticalLibraryIons = new List(); + peptide.Fragment(commonParameters.DissociationType, commonParameters.DigestionParams.FragmentationTerminus, theoreticalLibraryIons); + double mzCutoff = 300; + int fragmentNumberCutoff = 3; + + // if the spectrum has no peaks + if (scan.TheScan.MassSpectrum.XArray.Length == 0) + { + return 0; + } + List libraryDecoyIons = new List(); + Dictionary matchedIons = new Dictionary(); + + // search for each theoretical ion + for (int i = 0; i < theoreticalLibraryIons.Count; i++) + { + var libraryIon = theoreticalLibraryIons[i]; + double theoreticalFragmentMz = Math.Round(libraryIon.NeutralMass.ToMz(1) / 1.0005079, 0) * 1.0005079; + + // see https://www.nature.com/articles/s41592-019-0426-7 + // "All non-zero fragment ions (m/z > 300, ion >3, no neutral loss fragment ions) were considered for spectral angle calculation" + if (libraryIon.NeutralMass <= mzCutoff || libraryIon.FragmentNumber <= fragmentNumberCutoff) + { + continue; + } + + + + // get the closest peak in the spectrum to the library peak + var closestExperimentalMass = scan.GetClosestExperimentalIsotopicEnvelope(libraryIon.NeutralMass); + if (closestExperimentalMass != null && commonParameters.ProductMassTolerance.Within(closestExperimentalMass.MonoisotopicMass, libraryIon.NeutralMass) && closestExperimentalMass.Charge <= scan.PrecursorCharge)//TODO apply this filter before picking the envelope + { + var test = new Product(libraryIon.ProductType, libraryIon.Terminus, + libraryIon.NeutralMass, libraryIon.FragmentNumber, + libraryIon.AminoAcidPosition, libraryIon.NeutralLoss); + libraryDecoyIons.Add(new MatchedFragmentIon(ref libraryIon, theoreticalFragmentMz, 1, 1)); + matchedIons.Add(new MatchedFragmentIon(ref libraryIon, theoreticalFragmentMz, 1, 1), new MatchedFragmentIon(ref test, closestExperimentalMass.MonoisotopicMass.ToMz(closestExperimentalMass.Charge), closestExperimentalMass.Peaks.First().intensity, closestExperimentalMass.Charge)); + } + + } + + // L2 norm + double expNormalizer = Math.Sqrt(matchedIons.Sum(p => Math.Pow(p.Value.Intensity, 2))); + double theorNormalizer = Math.Sqrt(theoreticalLibraryIons.Sum(p => Math.Pow(1, 2))); + + double dotProduct = 0; + + foreach (var libraryIon in libraryDecoyIons) + { + if (matchedIons.TryGetValue(libraryIon, out var experIon)) + { + dotProduct += (libraryIon.Intensity / theorNormalizer) * (experIon.Intensity / expNormalizer); + } + } + + double normalizedSpectralAngle = 1 - (2 * Math.Acos(dotProduct) / Math.PI); + + return normalizedSpectralAngle; + } } } diff --git a/latest1129pepMTlibrary.csv b/latest1129pepMTlibrary.csv new file mode 100644 index 000000000..3b8f7c3f7 --- /dev/null +++ b/latest1129pepMTlibrary.csv @@ -0,0 +1,52015 @@ +PEP_QValue +0.000001 +0.000005 +0 +0.000002 +0.000001 +0.000002 +0.000002 +0.000002 +0.000001 +0.000002 +0.000003 +0.000002 +0.000002 +0 +0.000005 +0.000001 +0.000002 +0.000005 +0.000002 +0.000004 +0.000001 +0.000001 +0.000004 +0.000001 +0.000003 +0.000001 +0.000003 +0.000003 +0.000004 +0.000001 +0.000003 +0.000001 +0.000001 +0 +0.000003 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z1Ny(<*#;&D0MDN>{5KISFDC?Z{~!W*Gyerx2Ve#O_I9xHLfRVmcsaV+d;cB(zOt_z z(*S}K03QDO00n6Nfx*WC>53Hj$65665WJ1qaD4&LPYNKR{R;_ejQ=FTC(s4?w|(=l zz9Dgqp#YW1YlGPS0*VFyn{{soB+|zlUdg6 dNA{n<|1}`$YLEeZf Date: Thu, 3 Dec 2020 10:16:03 -0600 Subject: [PATCH 06/13] ClassicSearchEngine --- .../ClassicSearch/ClassicSearchEngine.cs | 126 +++++++++++++++++- .../SpectralLibrarySearchFunction.cs | 5 +- .../SearchTask/PostSearchAnalysisTask.cs | 2 +- 3 files changed, 124 insertions(+), 9 deletions(-) diff --git a/EngineLayer/ClassicSearch/ClassicSearchEngine.cs b/EngineLayer/ClassicSearch/ClassicSearchEngine.cs index 3200ceee4..1db4628bc 100644 --- a/EngineLayer/ClassicSearch/ClassicSearchEngine.cs +++ b/EngineLayer/ClassicSearch/ClassicSearchEngine.cs @@ -1,4 +1,5 @@ -using MzLibUtil; +using Chemistry; +using MzLibUtil; using Proteomics; using Proteomics.Fragmentation; using Proteomics.ProteolyticDigestion; @@ -112,10 +113,47 @@ protected override MetaMorpheusEngineResults RunSpecific() } } } - } - // report search progress (proteins searched so far out of total proteins in database) - proteinsSearched++; + List peptideDecoyProducts = GetDecoyProductsNB(peptideTheorProducts); + Protein decoyProtein = new Protein(peptide.Protein.BaseSequence, "DECOY_" + peptide.Protein.Accession, null, null, null, null, null, null, false, false, null, null, null, null, null, null, ""); + PeptideWithSetModifications decoyPeptide = new PeptideWithSetModifications(decoyProtein, new DigestionParams(), peptide.OneBasedStartResidueInProtein, peptide.OneBasedEndResidueInProtein, CleavageSpecificity.Full, peptide.PeptideDescription, peptide.MissedCleavages, new Dictionary(), peptide.NumFixedMods, null); + + foreach (ScanWithIndexAndNotchInfo scan in GetAcceptableScans(peptide.MonoisotopicMass, SearchMode)) + { + List matchedIons = MatchFragmentIons(scan.TheScan, peptideDecoyProducts, CommonParameters); + + double thisScore = CalculatePeptideScore(scan.TheScan.TheScan, matchedIons); + + bool meetsScoreCutoff = thisScore >= CommonParameters.ScoreCutoff; + + // this is thread-safe because even if the score improves from another thread writing to this PSM, + // the lock combined with AddOrReplace method will ensure thread safety + if (meetsScoreCutoff) + { + // valid hit (met the cutoff score); lock the scan to prevent other threads from accessing it + lock (myLocks[scan.ScanIndex]) + { + bool scoreImprovement = PeptideSpectralMatches[scan.ScanIndex] == null || (thisScore - PeptideSpectralMatches[scan.ScanIndex].RunnerUpScore) > -PeptideSpectralMatch.ToleranceForScoreDifferentiation; + + if (scoreImprovement) + { + if (PeptideSpectralMatches[scan.ScanIndex] == null) + { + PeptideSpectralMatches[scan.ScanIndex] = new PeptideSpectralMatch(decoyPeptide, scan.Notch, thisScore, scan.ScanIndex, scan.TheScan, CommonParameters.DigestionParams, matchedIons, 0); + } + else + { + PeptideSpectralMatches[scan.ScanIndex].AddOrReplace(decoyPeptide, thisScore, scan.Notch, CommonParameters.ReportAllAmbiguity, matchedIons, 0); + } + } + } + } + } + + } + + // report search progress (proteins searched so far out of total proteins in database) + proteinsSearched++; var percentProgress = (int)((proteinsSearched / Proteins.Count) * 100); if (percentProgress > oldPercentProgress) @@ -135,6 +173,86 @@ protected override MetaMorpheusEngineResults RunSpecific() return new MetaMorpheusEngineResults(this); } + private List GetDecoyProductsNB(List products) + { + List peptideTheorProducts = products.Where(p => p.TerminusFragment.Terminus == FragmentationTerminus.N).ToList(); + + List masses = peptideTheorProducts.Select(m => m.NeutralMass).ToList(); + masses = masses.OrderBy(m => m).ToList(); + List massDifferences = new List(); + for (int i = 0; i < masses.Count-1; i++) + { + massDifferences.Add(masses[i + 1] - masses[i]); + } + + massDifferences = ShuffleList(massDifferences); + + List newMasses = new List(); + newMasses.Add(masses.Max()); + double firstMass = masses.Max(); + foreach (double massDifference in massDifferences) + { + firstMass -= massDifference; + newMasses.Add(firstMass); + } + + List newProducts = new List(); + for (int i = 0; i < peptideTheorProducts.Count; i++) + { + NeutralTerminusFragment ntfOriginal = peptideTheorProducts[i].TerminusFragment; + NeutralTerminusFragment ntfNew = new NeutralTerminusFragment(ntfOriginal.Terminus, newMasses[i], ntfOriginal.FragmentNumber, ntfOriginal.AminoAcidPosition); + Product newProduct = new Product(peptideTheorProducts[i].ProductType, ntfNew, peptideTheorProducts[i].NeutralLoss); + newProducts.Add(newProduct); + } + + newProducts.AddRange(GetDecoyProductsCY(products.Where(p => p.TerminusFragment.Terminus == FragmentationTerminus.C).ToList(), massDifferences)); + + return newProducts; + } + + private List GetDecoyProductsCY(List peptideTheorProducts, List massDifferences) + { + List masses = peptideTheorProducts.Select(m => m.NeutralMass).ToList(); + masses = masses.OrderBy(m => m).ToList(); + + List newMasses = new List(); + + double firstMass = masses.Min() + ChemicalFormula.ParseFormula("H-2O-1").MonoisotopicMass; + newMasses.Add(firstMass); + + foreach (double massDifference in massDifferences) + { + firstMass += (massDifference); + newMasses.Add(firstMass); + } + + List newProducts = new List(); + for (int i = 0; i < peptideTheorProducts.Count; i++) + { + NeutralTerminusFragment ntfOriginal = peptideTheorProducts[i].TerminusFragment; + NeutralTerminusFragment ntfNew = new NeutralTerminusFragment(ntfOriginal.Terminus, newMasses[i], ntfOriginal.FragmentNumber, ntfOriginal.AminoAcidPosition); + Product newProduct = new Product(peptideTheorProducts[i].ProductType, ntfNew, peptideTheorProducts[i].NeutralLoss); + newProducts.Add(newProduct); + } + return newProducts; + } + + private List ShuffleList(List inputList) + { + List randomList = new List(); + + Random r = new Random(); + int randomIndex = 0; + while (inputList.Count > 0) + { + randomIndex = r.Next(0, inputList.Count); //Choose a random object in the list + randomList.Add(inputList[randomIndex]); //add it to the new, random list + inputList.RemoveAt(randomIndex); //remove to avoid duplicates + } + + return randomList; //return the new random list + } + private IEnumerable GetAcceptableScans(double peptideMonoisotopicMass, MassDiffAcceptor searchMode) { foreach (AllowedIntervalWithNotch allowedIntervalWithNotch in searchMode.GetAllowedPrecursorMassIntervalsFromTheoreticalMass(peptideMonoisotopicMass).ToList()) diff --git a/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs b/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs index 9174c5954..aa6c341db 100644 --- a/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs +++ b/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs @@ -33,10 +33,7 @@ public static void CalculateSpectralAngles(SpectralLibrary spectralLibrary, Pept double spectralAngle = CalculateNormalizedSpectralAngle(librarySpectrum.MatchedFragmentIons, scan.TheScan, commonParameters); psm.SpectralAngle = spectralAngle; } - else - { - psm.SpectralAngle = -1; - } + } } } diff --git a/TaskLayer/SearchTask/PostSearchAnalysisTask.cs b/TaskLayer/SearchTask/PostSearchAnalysisTask.cs index 506a56648..bcba2f9d3 100644 --- a/TaskLayer/SearchTask/PostSearchAnalysisTask.cs +++ b/TaskLayer/SearchTask/PostSearchAnalysisTask.cs @@ -521,7 +521,7 @@ private void SpectralLibraryGeneration() var FilteredPsmList = Parameters.AllPsms .Where(p => p.FdrInfo.QValue <= 0.1 && p.FdrInfo.QValueNotch <= CommonParameters.QValueOutputFilter).ToList(); - FilteredPsmList.RemoveAll(b => b.IsDecoy); + //FilteredPsmList.RemoveAll(b => b.IsDecoy); FilteredPsmList.RemoveAll(b => b.IsContaminant); //write spectral library by Yuling 11/14 From 69a9b3e9e99275f164f2e65da135548376110419 Mon Sep 17 00:00:00 2001 From: Yuling1996 Date: Wed, 16 Dec 2020 14:15:54 -0600 Subject: [PATCH 07/13] decoyUpdation --- .../ClassicSearch/ClassicSearchEngine.cs | 151 +++++----------- EngineLayer/DecoyOnTheFly.cs | 168 ++++++++++++++++++ EngineLayer/LibraryTargetDecoyDictionary.cs | 17 ++ EngineLayer/MetaMorpheusEngine.cs | 2 + EngineLayer/PeptideSpectralMatch.cs | 3 +- EngineLayer/ProteomicsExtensionMethods.cs | 38 ++++ .../SpectralLibrarySearch/LibrarySpectrum.cs | 6 +- .../SpectralLibrarySearch/SpectralLibrary.cs | 3 + .../SpectralLibrarySearchFunction.cs | 51 ++++-- TaskLayer/MetaMorpheusTask.cs | 57 +++--- .../SearchTask/PostSearchAnalysisTask.cs | 2 +- Test/SpectralLibraryReaderTest.cs | 40 ++++- 12 files changed, 376 insertions(+), 162 deletions(-) create mode 100644 EngineLayer/DecoyOnTheFly.cs create mode 100644 EngineLayer/LibraryTargetDecoyDictionary.cs create mode 100644 EngineLayer/ProteomicsExtensionMethods.cs diff --git a/EngineLayer/ClassicSearch/ClassicSearchEngine.cs b/EngineLayer/ClassicSearch/ClassicSearchEngine.cs index 1db4628bc..fcc13e086 100644 --- a/EngineLayer/ClassicSearch/ClassicSearchEngine.cs +++ b/EngineLayer/ClassicSearch/ClassicSearchEngine.cs @@ -67,6 +67,7 @@ protected override MetaMorpheusEngineResults RunSpecific() Parallel.ForEach(threads, (i) => { var peptideTheorProducts = new List(); + var decoyPeptideTheorProducts = new List(); for (; i < Proteins.Count; i += maxThreadsPerFile) { @@ -77,18 +78,29 @@ protected override MetaMorpheusEngineResults RunSpecific() foreach (PeptideWithSetModifications peptide in Proteins[i].Digest(CommonParameters.DigestionParams, FixedModifications, VariableModifications, SilacLabels, TurnoverLabels)) { peptide.Fragment(CommonParameters.DissociationType, CommonParameters.DigestionParams.FragmentationTerminus, peptideTheorProducts); + int[] newAAlocations = new int[peptide.BaseSequence.Length]; + PeptideWithSetModifications decoy = DecoyOnTheFly.GetReverseDecoyFromTarget(peptide, newAAlocations); + decoy.Fragment(CommonParameters.DissociationType, CommonParameters.DigestionParams.FragmentationTerminus, decoyPeptideTheorProducts); + foreach (ScanWithIndexAndNotchInfo scan in GetAcceptableScans(peptide.MonoisotopicMass, SearchMode)) { - //if (SpectralLibrary != null && !SpectralLibrary.ContainsSpectrum(peptide.FullSequence, scan.TheScan.PrecursorCharge)) - //{ - //continue; - //} - + if (SpectralLibrary != null && !SpectralLibrary.ContainsSpectrum(peptide.FullSequence, scan.TheScan.PrecursorCharge)) + { + continue; + } + // we need a function to get the original target sequence of a decoy peptide + if (SpectralLibrary != null && !SpectralLibrary.DecoyTargetPairs.ContainsKey(decoy)) + { + SpectralLibrary.DecoyTargetPairs.Add(decoy, peptide); + } + List matchedIons = MatchFragmentIons(scan.TheScan, peptideTheorProducts, CommonParameters); + List decoyMatchedIons = MatchFragmentIons(scan.TheScan, decoyPeptideTheorProducts, CommonParameters); double thisScore = CalculatePeptideScore(scan.TheScan.TheScan, matchedIons); - bool meetsScoreCutoff = thisScore >= CommonParameters.ScoreCutoff; + double decoyScore = CalculatePeptideScore(scan.TheScan.TheScan, decoyMatchedIons); + bool meetsScoreCutoff = Math.Max(thisScore, decoyScore) >= CommonParameters.ScoreCutoff; // this is thread-safe because even if the score improves from another thread writing to this PSM, // the lock combined with AddOrReplace method will ensure thread safety @@ -97,63 +109,45 @@ protected override MetaMorpheusEngineResults RunSpecific() // valid hit (met the cutoff score); lock the scan to prevent other threads from accessing it lock (myLocks[scan.ScanIndex]) { - bool scoreImprovement = PeptideSpectralMatches[scan.ScanIndex] == null || (thisScore - PeptideSpectralMatches[scan.ScanIndex].RunnerUpScore) > -PeptideSpectralMatch.ToleranceForScoreDifferentiation; + bool scoreImprovement = PeptideSpectralMatches[scan.ScanIndex] == null || (Math.Max(thisScore, decoyScore) - PeptideSpectralMatches[scan.ScanIndex].RunnerUpScore) > -PeptideSpectralMatch.ToleranceForScoreDifferentiation; if (scoreImprovement) { if (PeptideSpectralMatches[scan.ScanIndex] == null) { - PeptideSpectralMatches[scan.ScanIndex] = new PeptideSpectralMatch(peptide, scan.Notch, thisScore, scan.ScanIndex, scan.TheScan, CommonParameters, matchedIons, 0); + if (thisScore >= decoyScore) + { + PeptideSpectralMatches[scan.ScanIndex] = new PeptideSpectralMatch(peptide, scan.Notch, thisScore, scan.ScanIndex, scan.TheScan, CommonParameters, matchedIons, 0); + } + else + { + PeptideSpectralMatches[scan.ScanIndex] = new PeptideSpectralMatch(decoy, scan.Notch, decoyScore, scan.ScanIndex, scan.TheScan, CommonParameters, decoyMatchedIons, 0); + + } } else { - PeptideSpectralMatches[scan.ScanIndex].AddOrReplace(peptide, thisScore, scan.Notch, CommonParameters.ReportAllAmbiguity, matchedIons, 0); + if (thisScore >= decoyScore) + { + PeptideSpectralMatches[scan.ScanIndex].AddOrReplace(peptide, thisScore, scan.Notch, CommonParameters.ReportAllAmbiguity, matchedIons, 0); + } + else + { + PeptideSpectralMatches[scan.ScanIndex].AddOrReplace(decoy, decoyScore, scan.Notch, CommonParameters.ReportAllAmbiguity, decoyMatchedIons, 0); + } + } } - } - } - } - - List peptideDecoyProducts = GetDecoyProductsNB(peptideTheorProducts); - Protein decoyProtein = new Protein(peptide.Protein.BaseSequence, "DECOY_" + peptide.Protein.Accession, null, null, null, null, null, null, false, false, null, null, null, null, null, null, ""); - PeptideWithSetModifications decoyPeptide = new PeptideWithSetModifications(decoyProtein, new DigestionParams(), peptide.OneBasedStartResidueInProtein, peptide.OneBasedEndResidueInProtein, CleavageSpecificity.Full, peptide.PeptideDescription, peptide.MissedCleavages, new Dictionary(), peptide.NumFixedMods, null); - - foreach (ScanWithIndexAndNotchInfo scan in GetAcceptableScans(peptide.MonoisotopicMass, SearchMode)) - { - List matchedIons = MatchFragmentIons(scan.TheScan, peptideDecoyProducts, CommonParameters); - - double thisScore = CalculatePeptideScore(scan.TheScan.TheScan, matchedIons); - bool meetsScoreCutoff = thisScore >= CommonParameters.ScoreCutoff; - // this is thread-safe because even if the score improves from another thread writing to this PSM, - // the lock combined with AddOrReplace method will ensure thread safety - if (meetsScoreCutoff) - { - // valid hit (met the cutoff score); lock the scan to prevent other threads from accessing it - lock (myLocks[scan.ScanIndex]) - { - bool scoreImprovement = PeptideSpectralMatches[scan.ScanIndex] == null || (thisScore - PeptideSpectralMatches[scan.ScanIndex].RunnerUpScore) > -PeptideSpectralMatch.ToleranceForScoreDifferentiation; - - if (scoreImprovement) - { - if (PeptideSpectralMatches[scan.ScanIndex] == null) - { - PeptideSpectralMatches[scan.ScanIndex] = new PeptideSpectralMatch(decoyPeptide, scan.Notch, thisScore, scan.ScanIndex, scan.TheScan, CommonParameters.DigestionParams, matchedIons, 0); - } - else - { - PeptideSpectralMatches[scan.ScanIndex].AddOrReplace(decoyPeptide, thisScore, scan.Notch, CommonParameters.ReportAllAmbiguity, matchedIons, 0); - } - } } } + } + } - } - - // report search progress (proteins searched so far out of total proteins in database) - proteinsSearched++; + // report search progress (proteins searched so far out of total proteins in database) + proteinsSearched++; var percentProgress = (int)((proteinsSearched / Proteins.Count) * 100); if (percentProgress > oldPercentProgress) @@ -173,69 +167,6 @@ protected override MetaMorpheusEngineResults RunSpecific() return new MetaMorpheusEngineResults(this); } - private List GetDecoyProductsNB(List products) - { - List peptideTheorProducts = products.Where(p => p.TerminusFragment.Terminus == FragmentationTerminus.N).ToList(); - - List masses = peptideTheorProducts.Select(m => m.NeutralMass).ToList(); - masses = masses.OrderBy(m => m).ToList(); - List massDifferences = new List(); - for (int i = 0; i < masses.Count-1; i++) - { - massDifferences.Add(masses[i + 1] - masses[i]); - } - - massDifferences = ShuffleList(massDifferences); - - List newMasses = new List(); - newMasses.Add(masses.Max()); - double firstMass = masses.Max(); - foreach (double massDifference in massDifferences) - { - firstMass -= massDifference; - newMasses.Add(firstMass); - } - - List newProducts = new List(); - for (int i = 0; i < peptideTheorProducts.Count; i++) - { - NeutralTerminusFragment ntfOriginal = peptideTheorProducts[i].TerminusFragment; - NeutralTerminusFragment ntfNew = new NeutralTerminusFragment(ntfOriginal.Terminus, newMasses[i], ntfOriginal.FragmentNumber, ntfOriginal.AminoAcidPosition); - Product newProduct = new Product(peptideTheorProducts[i].ProductType, ntfNew, peptideTheorProducts[i].NeutralLoss); - newProducts.Add(newProduct); - } - - newProducts.AddRange(GetDecoyProductsCY(products.Where(p => p.TerminusFragment.Terminus == FragmentationTerminus.C).ToList(), massDifferences)); - - return newProducts; - } - - private List GetDecoyProductsCY(List peptideTheorProducts, List massDifferences) - { - List masses = peptideTheorProducts.Select(m => m.NeutralMass).ToList(); - masses = masses.OrderBy(m => m).ToList(); - - List newMasses = new List(); - - double firstMass = masses.Min() + ChemicalFormula.ParseFormula("H-2O-1").MonoisotopicMass; - newMasses.Add(firstMass); - - foreach (double massDifference in massDifferences) - { - firstMass += (massDifference); - newMasses.Add(firstMass); - } - - List newProducts = new List(); - for (int i = 0; i < peptideTheorProducts.Count; i++) - { - NeutralTerminusFragment ntfOriginal = peptideTheorProducts[i].TerminusFragment; - NeutralTerminusFragment ntfNew = new NeutralTerminusFragment(ntfOriginal.Terminus, newMasses[i], ntfOriginal.FragmentNumber, ntfOriginal.AminoAcidPosition); - Product newProduct = new Product(peptideTheorProducts[i].ProductType, ntfNew, peptideTheorProducts[i].NeutralLoss); - newProducts.Add(newProduct); - } - return newProducts; - } private List ShuffleList(List inputList) { diff --git a/EngineLayer/DecoyOnTheFly.cs b/EngineLayer/DecoyOnTheFly.cs new file mode 100644 index 000000000..1101c6095 --- /dev/null +++ b/EngineLayer/DecoyOnTheFly.cs @@ -0,0 +1,168 @@ +using Proteomics; +using Proteomics.ProteolyticDigestion; +using System; +using System.Collections.Generic; +using System.Linq; +using System.Text; + +namespace EngineLayer +{ + public class DecoyOnTheFly + { + //This function maintains the amino acids associated with the protease motif and reverses all other amino acids. + //N-terminal modificatons are preserved. Other modifications travel with their respective amino acids. this results + //in a decoy peptide composed the same amino acids and modifications as the original. + //Occasionally, this process results in peptide with exactly the same sequence. Therefore, there is a stop-gap measure + //the returns the mirror image of the original. N-terminal mods are preserved, but other mods are also reversed. + //this should yield a unique decoy for each target sequence. + public static PeptideWithSetModifications GetReverseDecoyFromTarget(PeptideWithSetModifications pwsm, int[] revisedAminoAcidOrder) + { + Dictionary newModificationsDictionary = new Dictionary(); + //Copy N-terminal modifications from target dictionary to decoy dictionary. + if (pwsm.AllModsOneIsNterminus.ContainsKey(1)) + { + newModificationsDictionary.Add(1, pwsm.AllModsOneIsNterminus[1]); + } + char[] newBase = new char[pwsm.BaseSequence.Length]; + ProteomicsExtenstionMethods.Fill(newBase, '0'); + char[] evaporatingBase = pwsm.BaseSequence.ToCharArray(); + List motifs = pwsm.DigestionParams.Protease.DigestionMotifs; + if (motifs != null && motifs.Count > 0) + { + foreach (var motif in motifs.Where(m => m.InducingCleavage != ""))//check the empty "" for topdown + { + string cleavingMotif = motif.InducingCleavage; + List cleavageMotifLocations = new List(); + + for (int i = 0; i < pwsm.BaseSequence.Length; i++) + { + bool Fits; + bool Prevents; + (Fits, Prevents) = motif.Fits(pwsm.BaseSequence, i); + + if (Fits && !Prevents) + { + cleavageMotifLocations.Add(i); + } + } + + foreach (int location in cleavageMotifLocations) + { + char[] motifArray = pwsm.BaseSequence.Substring(location, cleavingMotif.Length).ToCharArray(); + for (int i = 0; i < cleavingMotif.Length; i++) + { + newBase[location + i] = motifArray[i]; + revisedAminoAcidOrder[location + i] = location + i;// + //directly copy mods that were on amino acids in the motif. Those amino acids don't change position. + if (pwsm.AllModsOneIsNterminus.ContainsKey(location + i + 2)) + { + newModificationsDictionary.Add(location + i + 2, pwsm.AllModsOneIsNterminus[location + i + 2]); + } + + evaporatingBase[location + i] = '0';//can null a char so i use a number which doesnt' appear in peptide string + } + } + } + } + + //We've kept amino acids in the digestion motif in the same position in the decoy peptide. + //Now we will fill the remaining open positions in the decoy with the reverse of amino acids from the target. + int fillPosition = 0; + int extractPosition = pwsm.BaseSequence.Length - 1; + while (fillPosition < pwsm.BaseSequence.Length && extractPosition >= 0) + { + if (evaporatingBase[extractPosition] != '0') + { + while (newBase[fillPosition] != '0') + { + fillPosition++; + } + newBase[fillPosition] = evaporatingBase[extractPosition]; + revisedAminoAcidOrder[fillPosition] = extractPosition; + if (pwsm.AllModsOneIsNterminus.ContainsKey(extractPosition + 2)) + { + newModificationsDictionary.Add(fillPosition + 2, pwsm.AllModsOneIsNterminus[extractPosition + 2]); + } + fillPosition++; + } + extractPosition--; + } + + string newBaseString = new string(newBase); + + var proteinSequence = pwsm.Protein.BaseSequence; + var aStringBuilder = new StringBuilder(proteinSequence); + aStringBuilder.Remove(pwsm.OneBasedStartResidueInProtein - 1, pwsm.BaseSequence.Length); + aStringBuilder.Insert(pwsm.OneBasedStartResidueInProtein - 1, newBaseString); + proteinSequence = aStringBuilder.ToString(); + + Protein decoyProtein = new Protein(proteinSequence, "DECOY_" + pwsm.Protein.Accession, null, new List>(), new Dictionary>(), null, null, null, true); + + DigestionParams d = pwsm.DigestionParams; + + if (newBaseString != pwsm.BaseSequence) + { + return new PeptideWithSetModifications(decoyProtein, d, pwsm.OneBasedStartResidueInProtein, pwsm.OneBasedEndResidueInProtein, pwsm.CleavageSpecificityForFdrCategory, pwsm.PeptideDescription, pwsm.MissedCleavages, newModificationsDictionary, pwsm.NumFixedMods, newBaseString); + } + else + { + //The reverse decoy procedure failed to create a PeptideWithSetModificatons with a different sequence. Therefore, + //we retrun the mirror image peptide. + return GetPeptideMirror(pwsm, revisedAminoAcidOrder); + } + + } + + //Returns a PeptideWithSetModifications mirror image. Used when reverse decoy sequence is same as target sequence + public static PeptideWithSetModifications GetPeptideMirror(PeptideWithSetModifications pwsm, int[] revisedOrderNisOne) + { + Dictionary newModificationsDictionary = new Dictionary(); + //Copy N-terminal modifications from target dictionary to decoy dictionary. + if (pwsm.AllModsOneIsNterminus.ContainsKey(1)) + { + newModificationsDictionary.Add(1, pwsm.AllModsOneIsNterminus[1]); + } + + //First step is to reverse the position of all modifications except the mod on the peptide N-terminus. + if (pwsm.AllModsOneIsNterminus.Any()) + { + int newPosition = pwsm.BaseSequence.Length + 1; + for (int i = 2; i < pwsm.BaseSequence.Length + 2; i++) + { + if (pwsm.AllModsOneIsNterminus.ContainsKey(i)) + { + newModificationsDictionary.Add(newPosition, pwsm.AllModsOneIsNterminus[i]); + newPosition--; + } + else + { + newPosition--; + } + } + } + + //Second step is to reverse the sequence. + string newBaseString = ProteomicsExtenstionMethods.Reverse(pwsm.BaseSequence); + + var proteinSequence = pwsm.Protein.BaseSequence; + var aStringBuilder = new StringBuilder(proteinSequence); + aStringBuilder.Remove(pwsm.OneBasedStartResidueInProtein - 1, pwsm.BaseSequence.Length); + aStringBuilder.Insert(pwsm.OneBasedStartResidueInProtein - 1, newBaseString); + proteinSequence = aStringBuilder.ToString(); + + Protein decoyProtein = new Protein(proteinSequence, "DECOY_" + pwsm.Protein.Accession, null, new List>(), new Dictionary>(), null, null, null, true); + DigestionParams d = pwsm.DigestionParams; + + //now fill in the revised amino acid order + int oldStringPosition = pwsm.BaseSequence.Length - 1; + for (int i = 0; i < newBaseString.Length; i++) + { + revisedOrderNisOne[i] = oldStringPosition; + oldStringPosition--; + } + return new PeptideWithSetModifications(decoyProtein, d, pwsm.OneBasedStartResidueInProtein, pwsm.OneBasedEndResidueInProtein, pwsm.CleavageSpecificityForFdrCategory, pwsm.PeptideDescription, pwsm.MissedCleavages, newModificationsDictionary, pwsm.NumFixedMods, newBaseString); + } + + + } +} diff --git a/EngineLayer/LibraryTargetDecoyDictionary.cs b/EngineLayer/LibraryTargetDecoyDictionary.cs new file mode 100644 index 000000000..ca8764a96 --- /dev/null +++ b/EngineLayer/LibraryTargetDecoyDictionary.cs @@ -0,0 +1,17 @@ +using Proteomics.ProteolyticDigestion; +using System; +using System.Collections.Generic; +using System.Text; + +namespace EngineLayer +{ + public class LibraryTargetDecoyDictionary + { + public LibraryTargetDecoyDictionary() + { + + } + public Dictionary TargetDecoyPairs; + + } +} diff --git a/EngineLayer/MetaMorpheusEngine.cs b/EngineLayer/MetaMorpheusEngine.cs index 0d52ae8c9..356ccf0c9 100644 --- a/EngineLayer/MetaMorpheusEngine.cs +++ b/EngineLayer/MetaMorpheusEngine.cs @@ -138,6 +138,8 @@ public static List MatchFragmentIons(Ms2ScanWithSpecificMass { matchedFragmentIons.Add(new MatchedFragmentIon(ref product, closestExperimentalMass.MonoisotopicMass.ToMz(closestExperimentalMass.Charge), closestExperimentalMass.Peaks.First().intensity, closestExperimentalMass.Charge)); + var test1 = product.NeutralMass.ToMz(closestExperimentalMass.Charge); + var test2 = closestExperimentalMass.MonoisotopicMass.ToMz(closestExperimentalMass.Charge); } } if (commonParameters.AddCompIons) diff --git a/EngineLayer/PeptideSpectralMatch.cs b/EngineLayer/PeptideSpectralMatch.cs index 1bcb6160c..4122a6129 100644 --- a/EngineLayer/PeptideSpectralMatch.cs +++ b/EngineLayer/PeptideSpectralMatch.cs @@ -34,7 +34,8 @@ public PeptideSpectralMatch(PeptideWithSetModifications peptide, int notch, doub AddOrReplace(peptide, score, notch, true, matchedFragmentIons, xcorr); } - + + public string targetBeforeReverseTodecoy { get;set; } public ChemicalFormula ModsChemicalFormula { get; private set; } // these fields will be null if they are ambiguous public string FullSequence { get; private set; } public string EssentialSequence { get; private set; } diff --git a/EngineLayer/ProteomicsExtensionMethods.cs b/EngineLayer/ProteomicsExtensionMethods.cs new file mode 100644 index 000000000..db6803d77 --- /dev/null +++ b/EngineLayer/ProteomicsExtensionMethods.cs @@ -0,0 +1,38 @@ +using System; +using System.Collections.Generic; +using System.Text; + +namespace EngineLayer +{ + public static class ProteomicsExtenstionMethods + { + public static List AllIndexesOf(this string str, string value) + { + if (String.IsNullOrEmpty(value)) + throw new ArgumentException("the string to find may not be empty", "value"); + List indexes = new List(); + for (int index = 0; ; index += value.Length) + { + index = str.IndexOf(value, index); + if (index == -1) + return indexes; + indexes.Add(index); + } + } + + public static void Fill(this T[] originalArray, T with) + { + for (int i = 0; i < originalArray.Length; i++) + { + originalArray[i] = with; + } + } + + public static string Reverse(string s) + { + char[] charArray = s.ToCharArray(); + Array.Reverse(charArray); + return new string(charArray); + } + } +} diff --git a/EngineLayer/SpectralLibrarySearch/LibrarySpectrum.cs b/EngineLayer/SpectralLibrarySearch/LibrarySpectrum.cs index 3016df168..5108c5eb7 100644 --- a/EngineLayer/SpectralLibrarySearch/LibrarySpectrum.cs +++ b/EngineLayer/SpectralLibrarySearch/LibrarySpectrum.cs @@ -1,4 +1,5 @@ using Proteomics.Fragmentation; +using Proteomics.ProteolyticDigestion; using System.Collections.Generic; using System.Linq; using System.Text; @@ -19,6 +20,7 @@ public class LibrarySpectrum public bool IsDecoy { get; set; } public string ModsString { get; set; } public string BaseSequenceWithoutMods { get; set; } + public string Name { get { return Sequence + "/" + ChargeState; } @@ -50,7 +52,7 @@ public override string ToString() { double intensityFraction = matchedIon.Intensity / maxIntensity; - spectrum.Append("\n"+matchedIon.Mz + "\t" + intensityFraction + "\t" + "\"" + + spectrum.Append("\n" + matchedIon.Mz + "\t" + intensityFraction + "\t" + "\"" + matchedIon.NeutralTheoreticalProduct.ProductType.ToString() + matchedIon.NeutralTheoreticalProduct.FragmentNumber.ToString() + "^" + matchedIon.Charge + "/" + 0 + "ppm" + "\""); @@ -58,5 +60,7 @@ public override string ToString() return spectrum.ToString(); } + + } } \ No newline at end of file diff --git a/EngineLayer/SpectralLibrarySearch/SpectralLibrary.cs b/EngineLayer/SpectralLibrarySearch/SpectralLibrary.cs index 8f8b731b0..f9ff60862 100644 --- a/EngineLayer/SpectralLibrarySearch/SpectralLibrary.cs +++ b/EngineLayer/SpectralLibrarySearch/SpectralLibrary.cs @@ -1,5 +1,6 @@ using MzLibUtil; using Proteomics.Fragmentation; +using Proteomics.ProteolyticDigestion; using System; using System.Collections.Generic; using System.Globalization; @@ -19,6 +20,7 @@ public class SpectralLibrary private int MaxElementsInBuffer = 10000; private Dictionary StreamReaders; private static Regex IonParserRegex = new Regex(@"^(\D{1,})(\d{1,})(?:[\^]|$)(\d{1,}|$)"); + public Dictionary DecoyTargetPairs; private static Dictionary PrositToMetaMorpheusModDictionary = new Dictionary { @@ -33,6 +35,7 @@ public SpectralLibrary(List pathsToLibraries) LibrarySpectrumBufferList = new Queue(); LibrarySpectrumBuffer = new Dictionary(); StreamReaders = new Dictionary(); + DecoyTargetPairs = new Dictionary(); foreach (var path in LibraryPaths) { diff --git a/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs b/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs index aa6c341db..373ba6636 100644 --- a/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs +++ b/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs @@ -20,22 +20,53 @@ public static void CalculateSpectralAngles(SpectralLibrary spectralLibrary, Pept { Ms2ScanWithSpecificMass scan = arrayOfSortedMs2Scans[psm.ScanIndex]; - //TODO: spectral angle could be used to disambiguate PSMs. right now for ambiguous PSMs, the spectral angle for only one peptide option is saved - foreach (var peptide in psm.PeptidesToMatchingFragments) - { - if (spectralLibrary != null && peptide.Key.Protein.IsDecoy && !spectralLibrary.ContainsSpectrum(peptide.Key.FullSequence, scan.PrecursorCharge)) + //try + //{ + //TODO: spectral angle could be used to disambiguate PSMs. right now for ambiguous PSMs, the spectral angle for only one peptide option is saved + foreach (var peptide in psm.PeptidesToMatchingFragments) { - double spectralAngle = CalculateDecoyNormalizedSpectralAngle(peptide.Key, scan, commonParameters); - psm.SpectralAngle = spectralAngle; + if (spectralLibrary != null && !peptide.Key.Protein.Accession.Contains("DECOY") && spectralLibrary.TryGetSpectrum(peptide.Key.FullSequence, scan.PrecursorCharge, out var librarySpectrum)) + { + double spectralAngle = CalculateNormalizedSpectralAngle(librarySpectrum.MatchedFragmentIons, scan.TheScan, commonParameters); + psm.SpectralAngle = spectralAngle; + } + else if (spectralLibrary != null && peptide.Key.Protein.Accession.Contains("DECOY") && spectralLibrary.TryGetSpectrum(spectralLibrary.DecoyTargetPairs[peptide.Key].FullSequence, scan.PrecursorCharge, out var targetlibrarySpectrum)) + { + var decoyPeptideTheorProducts = new List(); + peptide.Key.Fragment(commonParameters.DissociationType, commonParameters.DigestionParams.FragmentationTerminus, decoyPeptideTheorProducts); + var decoylibrarySpectrum = GetDecoyLibrarySpectrumFromTargetByReverse(targetlibrarySpectrum, decoyPeptideTheorProducts); + double spectralAngle = CalculateNormalizedSpectralAngle(decoylibrarySpectrum, scan.TheScan, commonParameters); + psm.SpectralAngle = spectralAngle; + } } - else if (spectralLibrary != null && spectralLibrary.TryGetSpectrum(peptide.Key.FullSequence, scan.PrecursorCharge, out var librarySpectrum)) + //} + //catch + //{ + + //} + } + } + + public static List GetDecoyLibrarySpectrumFromTargetByReverse(LibrarySpectrum targetSpectrum, List decoyPeptideTheorProducts) + { + var decoyFragmentIons = new List(); + //var decoyPeptideTheorProducts = new List(); + //int[] newAAlocations = new int[targetPeptide.BaseSequence.Length]; + //PeptideWithSetModifications decoy = DecoyOnTheFly.GetReverseDecoyFromTarget(targetPeptide, newAAlocations); + //decoy.Fragment(CommonParameters.DissociationType, CommonParameters.DigestionParams.FragmentationTerminus, decoyPeptideTheorProducts); + foreach (var targetIon in targetSpectrum.MatchedFragmentIons) + { + foreach (var decoyPeptideTheorIon in decoyPeptideTheorProducts) + { + if (targetIon.NeutralTheoreticalProduct.ProductType == decoyPeptideTheorIon.ProductType && targetIon.NeutralTheoreticalProduct.FragmentNumber == decoyPeptideTheorIon.FragmentNumber) { - double spectralAngle = CalculateNormalizedSpectralAngle(librarySpectrum.MatchedFragmentIons, scan.TheScan, commonParameters); - psm.SpectralAngle = spectralAngle; + double decoyFragmentMz = decoyPeptideTheorIon.NeutralMass.ToMz(targetIon.Charge); + Product temProduct = decoyPeptideTheorIon; + decoyFragmentIons.Add(new MatchedFragmentIon(ref temProduct, decoyFragmentMz, targetIon.Intensity, targetIon.Charge)); } - } } + return decoyFragmentIons; } ///

diff --git a/TaskLayer/MetaMorpheusTask.cs b/TaskLayer/MetaMorpheusTask.cs index f1a172f1c..3b6a953c6 100644 --- a/TaskLayer/MetaMorpheusTask.cs +++ b/TaskLayer/MetaMorpheusTask.cs @@ -410,8 +410,8 @@ public MyTaskResults RunTask(string output_folder, List currentProtei FileSpecificParameters = new List<(string FileName, CommonParameters Parameters)>(); MetaMorpheusEngine.FinishedSingleEngineHandler += SingleEngineHandlerInTask; - try - { + //try + //{ var stopWatch = new Stopwatch(); stopWatch.Start(); @@ -458,25 +458,25 @@ public MyTaskResults RunTask(string output_folder, List currentProtei } FinishedWritingFile(resultsFileName, new List { displayName }); FinishedSingleTask(displayName); - } - catch (Exception e) - { - MetaMorpheusEngine.FinishedSingleEngineHandler -= SingleEngineHandlerInTask; - var resultsFileName = Path.Combine(output_folder, "results.txt"); - e.Data.Add("folder", output_folder); - using (StreamWriter file = new StreamWriter(resultsFileName)) - { - file.WriteLine(GlobalVariables.MetaMorpheusVersion.Equals("1.0.0.0") ? "MetaMorpheus: Not a release version" : "MetaMorpheus: version " + GlobalVariables.MetaMorpheusVersion); - file.WriteLine(SystemInfo.CompleteSystemInfo()); //OS, OS Version, .Net Version, RAM, processor count, MSFileReader .dll versions X3 - file.Write("e: " + e); - file.Write("e.Message: " + e.Message); - file.Write("e.InnerException: " + e.InnerException); - file.Write("e.Source: " + e.Source); - file.Write("e.StackTrace: " + e.StackTrace); - file.Write("e.TargetSite: " + e.TargetSite); - } - throw; - } + //} + //catch (Exception e) + //{ + // MetaMorpheusEngine.FinishedSingleEngineHandler -= SingleEngineHandlerInTask; + // var resultsFileName = Path.Combine(output_folder, "results.txt"); + // e.Data.Add("folder", output_folder); + // using (StreamWriter file = new StreamWriter(resultsFileName)) + // { + // file.WriteLine(GlobalVariables.MetaMorpheusVersion.Equals("1.0.0.0") ? "MetaMorpheus: Not a release version" : "MetaMorpheus: version " + GlobalVariables.MetaMorpheusVersion); + // file.WriteLine(SystemInfo.CompleteSystemInfo()); //OS, OS Version, .Net Version, RAM, processor count, MSFileReader .dll versions X3 + // file.Write("e: " + e); + // file.Write("e.Message: " + e.Message); + // file.Write("e.InnerException: " + e.InnerException); + // file.Write("e.Source: " + e.Source); + // file.Write("e.StackTrace: " + e.StackTrace); + // file.Write("e.TargetSite: " + e.TargetSite); + // } + // throw; + //} { var proseFilePath = Path.Combine(output_folder, "prose.txt"); @@ -597,21 +597,6 @@ protected static void WritePsmsToTsv(IEnumerable psms, str } } - protected static void WriteSpectralLibrary(List spectrumLibrary, string outputFolder) - { - - string spectrumFilePath = outputFolder + "\\spectralLibrary.msp"; - using (StreamWriter output = new StreamWriter(spectrumFilePath)) - { - - foreach (var x in spectrumLibrary) - { - output.WriteLine(x.ToString()); - } - - } - } - protected void ReportProgress(ProgressEventArgs v) { OutProgressHandler?.Invoke(this, v); diff --git a/TaskLayer/SearchTask/PostSearchAnalysisTask.cs b/TaskLayer/SearchTask/PostSearchAnalysisTask.cs index bcba2f9d3..6dfe6520c 100644 --- a/TaskLayer/SearchTask/PostSearchAnalysisTask.cs +++ b/TaskLayer/SearchTask/PostSearchAnalysisTask.cs @@ -563,7 +563,7 @@ private void SpectralLibraryGeneration() spectrumLibrary.Add(standSpectrum); } - WriteSpectralLibrary(spectrumLibrary, Parameters.OutputFolder); + //WriteSpectralLibrary(spectrumLibrary, Parameters.OutputFolder); } private void WritePsmResults() { diff --git a/Test/SpectralLibraryReaderTest.cs b/Test/SpectralLibraryReaderTest.cs index c2ebc519c..47f2d6a7a 100644 --- a/Test/SpectralLibraryReaderTest.cs +++ b/Test/SpectralLibraryReaderTest.cs @@ -7,6 +7,9 @@ using TaskLayer; using System.Collections.Generic; using Proteomics.Fragmentation; +using Proteomics; +using Proteomics.ProteolyticDigestion; +using Chemistry; namespace Test { @@ -158,7 +161,7 @@ public static void SpectralLibraryWriterTest() Assert.That(testLibraryWithoutDecoy.TryGetSpectrum("MVDENC[Common Fixed:Carbamidomethyl on C]VGFDHTVKPVSDMELETPTDK", 3, out var spectrum2)); } - [Test] + [Test] public static void TestSpectraCompareAndAverage() { var product = new Product(); @@ -170,7 +173,7 @@ public static void TestSpectraCompareAndAverage() var f = new MatchedFragmentIon(ref product, 2, 4, 1); var g = new MatchedFragmentIon(ref product, 3, 4, 1); var h = new MatchedFragmentIon(ref product, 5, 4, 1); - + var compare1 = SpectralLibrarySearchFunction.MatchedSpectraCompare(new List { a, b, c }, new List { f, g, h }); var compare2 = SpectralLibrarySearchFunction.MatchedSpectraCompare(new List { a, b, c }, new List { a, b, c }); var compare3 = SpectralLibrarySearchFunction.MatchedSpectraCompare(new List { a, b, h }, new List { f, e, g }); @@ -209,5 +212,36 @@ public static void TestSpectraCompareAndAverage() Assert.That(Math.Abs(ave2[1].Intensity - 0.3667) < 0.01); } - } + + [Test] + public static void TestDecoySpectralLibraryGeneration() + { + Product b1 = new Product(ProductType.b, FragmentationTerminus.Both, 1, 1, 0, 0); + Product b2 = new Product(ProductType.b, FragmentationTerminus.Both, 2, 2, 0, 0); + Product b3 = new Product(ProductType.b, FragmentationTerminus.Both, 3, 3, 0, 0); + Product b4 = new Product(ProductType.b, FragmentationTerminus.Both, 4, 4, 0, 0); + Product b5 = new Product(ProductType.b, FragmentationTerminus.Both, 5, 5, 0, 0); + Product y1 = new Product(ProductType.y, FragmentationTerminus.Both, 1, 1, 0, 0); + Product y2 = new Product(ProductType.y, FragmentationTerminus.Both, 2, 2, 0, 0); + Product y3 = new Product(ProductType.y, FragmentationTerminus.Both, 3, 3, 0, 0); + Product y4 = new Product(ProductType.y, FragmentationTerminus.Both, 4, 4, 0, 0); + Product y5 = new Product(ProductType.y, FragmentationTerminus.Both, 5, 5, 0, 0); + var o = new MatchedFragmentIon(ref b1, 1, 3, 1); + var p = new MatchedFragmentIon(ref b2, 2, 2, 2); + var q = new MatchedFragmentIon(ref b3, 3, 1, 1); + var r = new MatchedFragmentIon(ref y1, 4, 2, 2); + var s = new MatchedFragmentIon(ref y2, 2, 4, 1); + var t = new MatchedFragmentIon(ref y3, 2, 4, 2); + var test1LibrarySpectrum = new LibrarySpectrum("ABCD", 1, 3, + new List { o, p, q, r, s, t }, 0); + var test1TheoProducts = new List { b1, b2, b3, y1, y2, y3 }; + var decotTest1 = SpectralLibrarySearchFunction.GetDecoyLibrarySpectrumFromTargetByReverse(test1LibrarySpectrum, test1TheoProducts); + Assert.That(decotTest1[0].Intensity == 3); + Assert.That(decotTest1[1].Intensity == 2); + Assert.That(decotTest1[2].Intensity == 1); + Assert.That(decotTest1[1].Mz == b2.NeutralMass.ToMz(decotTest1[1].Charge)); + } + + +} } From 64cd3bb9a3bf7f8469e18d3eeaf549b61acf32e5 Mon Sep 17 00:00:00 2001 From: Yuling1996 Date: Thu, 17 Dec 2020 17:35:25 -0600 Subject: [PATCH 08/13] lastestWriter --- .../ClassicSearch/ClassicSearchEngine.cs | 4 +- .../SpectralLibrarySearch/LibrarySpectrum.cs | 29 ++++++++- .../SpectralLibrarySearchFunction.cs | 60 +++++++++++++++++-- TaskLayer/MetaMorpheusTask.cs | 28 +++++++++ .../SearchTask/PostSearchAnalysisTask.cs | 28 +++++++-- Test/SpectralLibraryReaderTest.cs | 12 ++-- 6 files changed, 144 insertions(+), 17 deletions(-) diff --git a/EngineLayer/ClassicSearch/ClassicSearchEngine.cs b/EngineLayer/ClassicSearch/ClassicSearchEngine.cs index fcc13e086..6115a1f2c 100644 --- a/EngineLayer/ClassicSearch/ClassicSearchEngine.cs +++ b/EngineLayer/ClassicSearch/ClassicSearchEngine.cs @@ -90,9 +90,11 @@ protected override MetaMorpheusEngineResults RunSpecific() continue; } // we need a function to get the original target sequence of a decoy peptide + if (SpectralLibrary != null && !SpectralLibrary.DecoyTargetPairs.ContainsKey(decoy)) { - SpectralLibrary.DecoyTargetPairs.Add(decoy, peptide); + + SpectralLibrary.DecoyTargetPairs.Add(decoy, peptide); } List matchedIons = MatchFragmentIons(scan.TheScan, peptideTheorProducts, CommonParameters); diff --git a/EngineLayer/SpectralLibrarySearch/LibrarySpectrum.cs b/EngineLayer/SpectralLibrarySearch/LibrarySpectrum.cs index 5108c5eb7..797990c00 100644 --- a/EngineLayer/SpectralLibrarySearch/LibrarySpectrum.cs +++ b/EngineLayer/SpectralLibrarySearch/LibrarySpectrum.cs @@ -36,6 +36,32 @@ public LibrarySpectrum(string sequence, double precursorMz, int chargeState, Lis RetentionTime = rt; } + //public override string ToString() + //{ + // StringBuilder spectrum = new StringBuilder(); + // spectrum.Append("Name: " + Name); + // spectrum.Append("\nMW: " + PrecursorMz); + // spectrum.Append("\nComment: "); + // spectrum.Append("Parent=" + PrecursorMz); + // spectrum.Append(" RT=" + RetentionTime); + // spectrum.Append("\nNum peaks: " + MatchedFragmentIons.Count); + + // double maxIntensity = MatchedFragmentIons.Select(b => b.Intensity).Max(); + + // foreach (MatchedFragmentIon matchedIon in MatchedFragmentIons) + // { + // double intensityFraction = matchedIon.Intensity / maxIntensity; + + // spectrum.Append("\n" + matchedIon.Mz + "\t" + intensityFraction + "\t" + "\"" + + // matchedIon.NeutralTheoreticalProduct.ProductType.ToString() + + // matchedIon.NeutralTheoreticalProduct.FragmentNumber.ToString() + "^" + + // matchedIon.Charge + "/" + 0 + "ppm" + "\""); + // } + + // return spectrum.ToString(); + //} + + //test public override string ToString() { StringBuilder spectrum = new StringBuilder(); @@ -47,10 +73,11 @@ public override string ToString() spectrum.Append("\nNum peaks: " + MatchedFragmentIons.Count); double maxIntensity = MatchedFragmentIons.Select(b => b.Intensity).Max(); + double sumIntensity = MatchedFragmentIons.Select(b => b.Intensity).Sum(); foreach (MatchedFragmentIon matchedIon in MatchedFragmentIons) { - double intensityFraction = matchedIon.Intensity / maxIntensity; + double intensityFraction = matchedIon.Intensity / sumIntensity; spectrum.Append("\n" + matchedIon.Mz + "\t" + intensityFraction + "\t" + "\"" + matchedIon.NeutralTheoreticalProduct.ProductType.ToString() + diff --git a/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs b/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs index 373ba6636..bc97be660 100644 --- a/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs +++ b/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs @@ -132,18 +132,64 @@ public static double CalculateNormalizedSpectralAngle(List t return normalizedSpectralAngle; } + public static double CalculatePsmsNormalizedSpectralAngle(List standardSpectrum, List spectrum) + { + double mzCutoff = 300; + int fragmentNumberCutoff = 3; + Dictionary matchedIons = new Dictionary(); + foreach (var standIon in standardSpectrum) + { + if (standIon.Mz <= mzCutoff || standIon.NeutralTheoreticalProduct.FragmentNumber <= fragmentNumberCutoff) + { + continue; + } + foreach (var ion in spectrum) + { + if (standIon.NeutralTheoreticalProduct.ProductType == ion.NeutralTheoreticalProduct.ProductType && standIon.NeutralTheoreticalProduct.FragmentNumber == ion.NeutralTheoreticalProduct.FragmentNumber && standIon.Charge == ion.Charge) + { + matchedIons.Add(standIon, ion); + } + } + } + + // L2 norm + double sumOfspectrumIntensity = Math.Sqrt(matchedIons.Sum(p => p.Value.Intensity)); + double expNormalizer = Math.Sqrt(matchedIons.Sum(p => Math.Pow(p.Value.Intensity / sumOfspectrumIntensity, 2))); + double sumOfstandardIntensity = Math.Sqrt(standardSpectrum.Sum(p => p.Intensity)); + double theorNormalizer = Math.Sqrt(standardSpectrum.Sum(p => Math.Pow(p.Intensity/ sumOfstandardIntensity, 2))); + + double dotProduct = 0; + + foreach (var standIon in standardSpectrum) + { + if (matchedIons.TryGetValue(standIon, out var Ion)) + { + var standIntensity = standIon.Intensity / sumOfstandardIntensity; + var intensity = Ion.Intensity / sumOfspectrumIntensity; + dotProduct += (standIntensity / theorNormalizer) * (intensity / expNormalizer); + } + } + + double normalizedSpectralAngle = 1 - (2 * Math.Acos(dotProduct) / Math.PI); + + return normalizedSpectralAngle; + + } + //1024TestDoneCompareFunction public static double MatchedSpectraCompare(List standardSpectra, List spectraToCompare) { double[] mz1 = standardSpectra.Select(b => b.Mz).ToArray(); double intensitySum1 = standardSpectra.Select(b => b.Intensity).Sum(); + //sqrt of the intensity to decrease the influence of peaks with high intensity double[] intensity1 = standardSpectra.Select(b => Math.Sqrt(b.Intensity / intensitySum1)).ToArray(); //Console.WriteLine(mz1.Length + " " + intensity1.Length); Array.Sort(mz1, intensity1); double[] mz2 = spectraToCompare.Select(b => b.Mz).ToArray(); double intensitySum2 = spectraToCompare.Select(b => b.Intensity).Sum(); + //sqrt of the intensity to decrease the influence of peaks with high intensity double[] intensity2 = spectraToCompare.Select(b => Math.Sqrt(b.Intensity / intensitySum2)).ToArray(); Array.Sort(mz2, intensity2); //Console.WriteLine(mz2.Length + " " + intensity2.Length); @@ -216,7 +262,7 @@ public static double MatchedSpectraCompare(List standardSpec return score; } - public static List AverageTwoSpectra(List spectraOne, List spectraTwo) + public static List AverageTwoSpectra(List spectraOne, List spectraTwo, int PsmsNum) { Dictionary averagedPeaksDictionary = new Dictionary(); var averageTwoSpectraResult = new List(); @@ -252,8 +298,10 @@ public static List AverageTwoSpectra(List AverageTwoSpectra(List psms, str } } } + protected static void WriteSpectralLibrary(List spectrumLibrary, string outputFolder) + { + + string spectrumFilePath = outputFolder + "\\spectralLibrary.msp"; + using (StreamWriter output = new StreamWriter(spectrumFilePath)) + { + foreach (var x in spectrumLibrary) + { + output.WriteLine(x.ToString()); + } + + } + } + //forTestingLibraryGeneration + protected static void WriteAllspectra(List spectrumLibrary, string outputFolder) + { + + string spectrumFilePath = outputFolder + "\\allspectral.msp"; + using (StreamWriter output = new StreamWriter(spectrumFilePath)) + { + + foreach (var x in spectrumLibrary) + { + output.WriteLine(x.ToString()); + } + + } + } protected void ReportProgress(ProgressEventArgs v) { OutProgressHandler?.Invoke(this, v); diff --git a/TaskLayer/SearchTask/PostSearchAnalysisTask.cs b/TaskLayer/SearchTask/PostSearchAnalysisTask.cs index 6dfe6520c..4747282c5 100644 --- a/TaskLayer/SearchTask/PostSearchAnalysisTask.cs +++ b/TaskLayer/SearchTask/PostSearchAnalysisTask.cs @@ -521,7 +521,8 @@ private void SpectralLibraryGeneration() var FilteredPsmList = Parameters.AllPsms .Where(p => p.FdrInfo.QValue <= 0.1 && p.FdrInfo.QValueNotch <= CommonParameters.QValueOutputFilter).ToList(); - //FilteredPsmList.RemoveAll(b => b.IsDecoy); + FilteredPsmList.RemoveAll(b => b.BestMatchingPeptides.First().Peptide.Protein.Accession.Contains("DECOY")); + FilteredPsmList.RemoveAll(b => b.IsDecoy); FilteredPsmList.RemoveAll(b => b.IsContaminant); //write spectral library by Yuling 11/14 @@ -544,17 +545,21 @@ private void SpectralLibraryGeneration() double standPrecursurMz = x.Value[0].ScanPrecursorMonoisotopicPeakMz; double standRt = x.Value[0].ScanRetentionTime; int a = 0; + int PsmsNum = 1; while (a < x.Value.Count - 1) { //Console.WriteLine(x.Key + " " + this.matchedSpectraCompare(standspctra, x.Value[a + 1].MatchedFragmentIons)); var spectrumTocompare = x.Value[a + 1].MatchedFragmentIons; var compareScore = SpectralLibrarySearchFunction.MatchedSpectraCompare(standspctra, spectrumTocompare); + var testScore = SpectralLibrarySearchFunction.CalculatePsmsNormalizedSpectralAngle(standspctra, spectrumTocompare); if (compareScore > 0.90) { - standspctra = SpectralLibrarySearchFunction.AverageTwoSpectra(standspctra, spectrumTocompare); - standPrecursurMz = (standPrecursurMz + x.Value[a + 1].ScanPrecursorMonoisotopicPeakMz) / 2; - standRt = (standRt + x.Value[a + 1].ScanRetentionTime) / 2; + PsmsNum++; + standspctra = SpectralLibrarySearchFunction.AverageTwoSpectra(standspctra, spectrumTocompare, PsmsNum); + + standPrecursurMz = standPrecursurMz * (1 - (1.0 / PsmsNum)) + x.Value[a + 1].ScanPrecursorMonoisotopicPeakMz / PsmsNum; + standRt = standRt * (1 - (1.0 / PsmsNum)) + x.Value[a + 1].ScanRetentionTime / PsmsNum; } a++; } @@ -564,6 +569,21 @@ private void SpectralLibraryGeneration() } //WriteSpectralLibrary(spectrumLibrary, Parameters.OutputFolder); + + //fortesting + var allspectra = new List(); + foreach (var x in PsmsGroupByPeptideAndCharge) + { + if(x.Value.Count>1) + { + foreach(var each in x.Value) + { + allspectra.Add(new LibrarySpectrum(each.FullSequence, each.ScanPrecursorMonoisotopicPeakMz, each.ScanPrecursorCharge, each.MatchedFragmentIons, each.ScanRetentionTime)); + } + } + + } + WriteAllspectra(allspectra, Parameters.OutputFolder); } private void WritePsmResults() { diff --git a/Test/SpectralLibraryReaderTest.cs b/Test/SpectralLibraryReaderTest.cs index 47f2d6a7a..35a30505f 100644 --- a/Test/SpectralLibraryReaderTest.cs +++ b/Test/SpectralLibraryReaderTest.cs @@ -177,9 +177,9 @@ public static void TestSpectraCompareAndAverage() var compare1 = SpectralLibrarySearchFunction.MatchedSpectraCompare(new List { a, b, c }, new List { f, g, h }); var compare2 = SpectralLibrarySearchFunction.MatchedSpectraCompare(new List { a, b, c }, new List { a, b, c }); var compare3 = SpectralLibrarySearchFunction.MatchedSpectraCompare(new List { a, b, h }, new List { f, e, g }); - Assert.That(Math.Abs(compare1 - 0.6667) < 0.001); + Assert.That(Math.Abs(compare1-0.6667)<0.001); Assert.That(Math.Abs(compare2 - 1) < 0.001); - Assert.That(Math.Abs(compare3 - 0.4714) < 0.001); + Assert.That(Math.Abs(compare3-0.5774)<0.001); //test averageTwoSpectraFunction by Yuling Product b1 = new Product(ProductType.b, FragmentationTerminus.Both, 0, 1, 0, 0); @@ -201,15 +201,15 @@ public static void TestSpectraCompareAndAverage() var t = new MatchedFragmentIon(ref b3, 2, 4, 2); // all same charge - var ave1 = SpectralLibrarySearchFunction.AverageTwoSpectra(new List { o, p, q }, new List { o, p, q }); + var ave1 = SpectralLibrarySearchFunction.AverageTwoSpectra(new List { o, p, q }, new List { o, p, q },2); Assert.AreEqual(ave1.Count, 3); Assert.AreEqual(ave1[0], new MatchedFragmentIon(ref b1, 1, 0.5, 1)); // with different charges - var ave2 = SpectralLibrarySearchFunction.AverageTwoSpectra(new List { o, p, q }, new List { r, s, t }); + var ave2 = SpectralLibrarySearchFunction.AverageTwoSpectra(new List { o, p, q }, new List { r, s, t },3); Assert.AreEqual(ave2.Count, 5); - Assert.AreEqual(ave2[0].Intensity, 0.25); - Assert.That(Math.Abs(ave2[1].Intensity - 0.3667) < 0.01); + Assert.That(Math.Abs(ave2[0].Intensity-0.33333)< 0.001); + Assert.That(Math.Abs(ave2[1].Intensity - 0.35556) < 0.001); } From 7993b599f12dc3ee3c53b4a38c7638f90d5e837e Mon Sep 17 00:00:00 2001 From: Yuling1996 Date: Wed, 13 Jan 2021 14:56:57 -0600 Subject: [PATCH 09/13] troubleShooting --- .../ClassicSearch/ClassicSearchEngine.cs | 37 ++-- .../FdrAnalysis/PEPValueAnalysisGeneric.cs | 2 +- EngineLayer/PeptideSpectralMatch.cs | 2 + .../SpectralLibrarySearch/LibrarySpectrum.cs | 3 +- .../SpectralLibrarySearch/SpectralLibrary.cs | 163 +++++++++++++++++- .../SpectralLibrarySearchFunction.cs | 55 ++++-- .../SearchTask/PostSearchAnalysisTask.cs | 46 +++-- Test/SpectralLibraryReaderTest.cs | 56 ++++-- Test/Test.csproj | 6 +- Test/TestData/yeast2fake_pdeep_lib.msp | 71 ++++++++ 10 files changed, 373 insertions(+), 68 deletions(-) create mode 100644 Test/TestData/yeast2fake_pdeep_lib.msp diff --git a/EngineLayer/ClassicSearch/ClassicSearchEngine.cs b/EngineLayer/ClassicSearch/ClassicSearchEngine.cs index 6115a1f2c..94a701559 100644 --- a/EngineLayer/ClassicSearch/ClassicSearchEngine.cs +++ b/EngineLayer/ClassicSearch/ClassicSearchEngine.cs @@ -12,7 +12,8 @@ namespace EngineLayer.ClassicSearch { public class ClassicSearchEngine : MetaMorpheusEngine { - private readonly SpectralLibrary SpectralLibrary; + public SpectralLibrary SpectralLibrary { get; set; } + public Dictionary decoyTargetPairs; private readonly MassDiffAcceptor SearchMode; private readonly List Proteins; private readonly List FixedModifications; @@ -50,9 +51,10 @@ protected override MetaMorpheusEngineResults RunSpecific() double proteinsSearched = 0; int oldPercentProgress = 0; + decoyTargetPairs = new Dictionary(); - // one lock for each MS2 scan; a scan can only be accessed by one thread at a time - var myLocks = new object[PeptideSpectralMatches.Length]; + // one lock for each MS2 scan; a scan can only be accessed by one thread at a time + var myLocks = new object[PeptideSpectralMatches.Length]; for (int i = 0; i < myLocks.Length; i++) { myLocks[i] = new object(); @@ -82,21 +84,23 @@ protected override MetaMorpheusEngineResults RunSpecific() PeptideWithSetModifications decoy = DecoyOnTheFly.GetReverseDecoyFromTarget(peptide, newAAlocations); decoy.Fragment(CommonParameters.DissociationType, CommonParameters.DigestionParams.FragmentationTerminus, decoyPeptideTheorProducts); + // we need a function to get the original target sequence of a decoy peptide - foreach (ScanWithIndexAndNotchInfo scan in GetAcceptableScans(peptide.MonoisotopicMass, SearchMode)) - { - if (SpectralLibrary != null && !SpectralLibrary.ContainsSpectrum(peptide.FullSequence, scan.TheScan.PrecursorCharge)) + if (!decoyTargetPairs.ContainsKey(decoy) ) { - continue; + decoyTargetPairs.Add(decoy, peptide); } - // we need a function to get the original target sequence of a decoy peptide + - if (SpectralLibrary != null && !SpectralLibrary.DecoyTargetPairs.ContainsKey(decoy)) - { - - SpectralLibrary.DecoyTargetPairs.Add(decoy, peptide); - } - + + + foreach (ScanWithIndexAndNotchInfo scan in GetAcceptableScans(peptide.MonoisotopicMass, SearchMode)) + { + //if (SpectralLibrary != null && !SpectralLibrary.ContainsSpectrum(peptide.FullSequence, scan.TheScan.PrecursorCharge)) + //{ + // continue; + //} + List matchedIons = MatchFragmentIons(scan.TheScan, peptideTheorProducts, CommonParameters); List decoyMatchedIons = MatchFragmentIons(scan.TheScan, decoyPeptideTheorProducts, CommonParameters); @@ -147,7 +151,6 @@ protected override MetaMorpheusEngineResults RunSpecific() } } - // report search progress (proteins searched so far out of total proteins in database) proteinsSearched++; var percentProgress = (int)((proteinsSearched / Proteins.Count) * 100); @@ -165,6 +168,10 @@ protected override MetaMorpheusEngineResults RunSpecific() { psm.ResolveAllAmbiguities(); } + if (SpectralLibrary != null) + { + SpectralLibrary.DecoyTargetPairs = decoyTargetPairs; + } return new MetaMorpheusEngineResults(this); } diff --git a/EngineLayer/FdrAnalysis/PEPValueAnalysisGeneric.cs b/EngineLayer/FdrAnalysis/PEPValueAnalysisGeneric.cs index 154a0ab10..63170791d 100644 --- a/EngineLayer/FdrAnalysis/PEPValueAnalysisGeneric.cs +++ b/EngineLayer/FdrAnalysis/PEPValueAnalysisGeneric.cs @@ -625,7 +625,7 @@ public static PsmData CreateOnePsmDataEntry(string searchType, List<(string file float isLoop = 0; float isInter = 0; float isIntra = 0; - float spectralAngle = 0; + float spectralAngle = (float)double.NaN; if (searchType != "crosslink") { diff --git a/EngineLayer/PeptideSpectralMatch.cs b/EngineLayer/PeptideSpectralMatch.cs index 4122a6129..5efa8f5e9 100644 --- a/EngineLayer/PeptideSpectralMatch.cs +++ b/EngineLayer/PeptideSpectralMatch.cs @@ -3,6 +3,7 @@ using Proteomics; using Proteomics.Fragmentation; using Proteomics.ProteolyticDigestion; +using System; using System.Collections.Generic; using System.Linq; @@ -31,6 +32,7 @@ public PeptideSpectralMatch(PeptideWithSetModifications peptide, int notch, doub Xcorr = xcorr; NativeId = scan.NativeId; RunnerUpScore = commonParameters.ScoreCutoff; + SpectralAngle = Double.NaN; AddOrReplace(peptide, score, notch, true, matchedFragmentIons, xcorr); } diff --git a/EngineLayer/SpectralLibrarySearch/LibrarySpectrum.cs b/EngineLayer/SpectralLibrarySearch/LibrarySpectrum.cs index 797990c00..4451a6351 100644 --- a/EngineLayer/SpectralLibrarySearch/LibrarySpectrum.cs +++ b/EngineLayer/SpectralLibrarySearch/LibrarySpectrum.cs @@ -77,7 +77,8 @@ public override string ToString() foreach (MatchedFragmentIon matchedIon in MatchedFragmentIons) { - double intensityFraction = matchedIon.Intensity / sumIntensity; + //double intensityFraction = matchedIon.Intensity / sumIntensity; + double intensityFraction = matchedIon.Intensity / maxIntensity; spectrum.Append("\n" + matchedIon.Mz + "\t" + intensityFraction + "\t" + "\"" + matchedIon.NeutralTheoreticalProduct.ProductType.ToString() + diff --git a/EngineLayer/SpectralLibrarySearch/SpectralLibrary.cs b/EngineLayer/SpectralLibrarySearch/SpectralLibrary.cs index f9ff60862..14fad55a9 100644 --- a/EngineLayer/SpectralLibrarySearch/SpectralLibrary.cs +++ b/EngineLayer/SpectralLibrarySearch/SpectralLibrary.cs @@ -20,7 +20,7 @@ public class SpectralLibrary private int MaxElementsInBuffer = 10000; private Dictionary StreamReaders; private static Regex IonParserRegex = new Regex(@"^(\D{1,})(\d{1,})(?:[\^]|$)(\d{1,}|$)"); - public Dictionary DecoyTargetPairs; + public Dictionary DecoyTargetPairs { get; set; } private static Dictionary PrositToMetaMorpheusModDictionary = new Dictionary { @@ -28,6 +28,12 @@ public class SpectralLibrary { "Carbamidomethyl", "[Common Fixed:Carbamidomethyl on C]" } }; + private static Dictionary pDeepToMetaMorpheusModDictionary = new Dictionary + { + { "Oxidation","[Common Variable:Oxidation on M]" }, + {"CAM", "[Common Fixed:Carbamidomethyl on C]" } + }; + public SpectralLibrary(List pathsToLibraries) { LibraryPaths = pathsToLibraries; @@ -152,7 +158,15 @@ private LibrarySpectrum ReadSpectrumFromLibraryFile(string path, long byteOffset reader.DiscardBufferedData(); // return the library spectrum - return ReadLibrarySpectrum(reader); + if(path.Contains("pdeep")) + { + return ReadLibrarySpectrum_pDeep(reader); + } + else + { + return ReadLibrarySpectrum(reader); + } + } private LibrarySpectrum ReadLibrarySpectrum(StreamReader reader, bool onlyReadHeader = false) @@ -351,6 +365,140 @@ private LibrarySpectrum ReadLibrarySpectrum(StreamReader reader, bool onlyReadHe return new LibrarySpectrum(sequence, precursorMz, z, matchedFragmentIons, rt); } + private LibrarySpectrum ReadLibrarySpectrum_pDeep(StreamReader reader, bool onlyReadHeader = false) + { + char[] nameSplit = new char[] { '/', '_' }; + char[] mwSplit = new char[] { ':' }; + char[] commentSplit = new char[] { ' ', ':', '=' }; + char[] modSplit = new char[] { '/', '(', ')' }; + char[] fragmentSplit = new char[] { '\t', '/' }; + + bool readingPeaks = false; + string sequence = null; + int z = 2; + double precursorMz = 0; + double rt = 0; + List matchedFragmentIons = new List(); + + while (reader.Peek() > 0) + { + string line = reader.ReadLine(); + string[] split; + + if (line.StartsWith("Name", StringComparison.InvariantCultureIgnoreCase)) + { + if (sequence != null) + { + return new LibrarySpectrum(sequence, precursorMz, z, matchedFragmentIons, rt); + } + + split = line.Split(nameSplit); + + // get sequence + sequence = split[0].Replace("Name:", string.Empty).Trim(); + + // get charge + z = int.Parse(split[1].Trim()); + + string[] mods = split[2].Split(modSplit, StringSplitOptions.RemoveEmptyEntries); + for (int i = mods.Length - 1; i > 0; i--) + { + string[] modInfo = mods[i].Split(','); + int index = Convert.ToInt32(modInfo[0]); + string mod = modInfo[2]; + string metaMorpheusMod = pDeepToMetaMorpheusModDictionary[mod]; + //add the mod into the sequence + string leftSeq = sequence.Substring(0, index + 1); + string rightSeq = sequence.Substring(index + 1); + sequence = leftSeq + metaMorpheusMod + rightSeq; + } + + } + else if (line.StartsWith("MW", StringComparison.InvariantCultureIgnoreCase)) + { + split = line.Split(mwSplit); + + // get precursor m/z + precursorMz = double.Parse(split[1].Trim(), CultureInfo.InvariantCulture); + } + else if (line.StartsWith("Comment", StringComparison.InvariantCultureIgnoreCase)) + { + split = line.Split(commentSplit); + + // get precursor m/z if not defined yet + if (precursorMz == 0) + { + int indOfParent = Array.IndexOf(split, "Parent"); + if (indOfParent > 0) + { + precursorMz = double.Parse(split[indOfParent + 1]); + } + } + + // get RT + int indOfRt = Array.IndexOf(split, "iRT"); + if (indOfRt > 0) + { + rt = double.Parse(split[indOfRt + 1]); + } + else + { + indOfRt = Array.IndexOf(split, "RT"); + + if (indOfRt > 0) + { + rt = double.Parse(split[indOfRt + 1]); + } + } + } + else if (line.StartsWith("Num peaks", StringComparison.InvariantCultureIgnoreCase)) + { + if (onlyReadHeader) + { + return new LibrarySpectrum(sequence, precursorMz, z, matchedFragmentIons, rt); + } + + // this assumes that the peaks are listed after the "Num peaks" line + readingPeaks = true; + } + else if (readingPeaks && line != "") + { + split = line.Split(fragmentSplit, StringSplitOptions.RemoveEmptyEntries); + + // read fragment m/z + var experMz = double.Parse(split[0], CultureInfo.InvariantCulture); + + // read fragment intensity + var experIntensity = double.Parse(split[1], CultureInfo.InvariantCulture); + + // read fragment type, number + + string fragmentType = split[2].ToCharArray()[0].ToString(); + int fragmentNumber = int.Parse(new string(split[2].Split(new char[] { '^' })[0].Where(Char.IsDigit).ToArray())); + int fragmentCharge = 1; + + + if (split[2].Contains('^')) + { + fragmentCharge = int.Parse(split[2].Split('^')[1]); + } + + ProductType peakProductType = (ProductType)Enum.Parse(typeof(ProductType), fragmentType, true); + + //TODO: figure out terminus + FragmentationTerminus terminus = (FragmentationTerminus)Enum.Parse(typeof(FragmentationTerminus), "None", true); + + //TODO: figure out amino acid position + var product = new Product(peakProductType, terminus, experMz, fragmentNumber, 0, 0); + + matchedFragmentIons.Add(new MatchedFragmentIon(ref product, experMz, experIntensity, fragmentCharge)); + } + } + + return new LibrarySpectrum(sequence, precursorMz, z, matchedFragmentIons, rt); + } + + private void IndexSpectralLibrary(string path) { var reader = new StreamReader(path); @@ -371,7 +519,16 @@ private void IndexSpectralLibrary(string path) reader.DiscardBufferedData(); // parse the header - var libraryItem = ReadLibrarySpectrum(reader, onlyReadHeader: true); + LibrarySpectrum libraryItem; + if (path.Contains("pdeep")) + { + libraryItem = ReadLibrarySpectrum_pDeep(reader, onlyReadHeader: true); + } + else + { + libraryItem = ReadLibrarySpectrum(reader, onlyReadHeader: true); + } + // add the spectrum to the index SequenceToFileAndLocation.TryAdd(libraryItem.Name, (path, byteOffset)); diff --git a/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs b/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs index bc97be660..3c83331da 100644 --- a/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs +++ b/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs @@ -30,7 +30,7 @@ public static void CalculateSpectralAngles(SpectralLibrary spectralLibrary, Pept double spectralAngle = CalculateNormalizedSpectralAngle(librarySpectrum.MatchedFragmentIons, scan.TheScan, commonParameters); psm.SpectralAngle = spectralAngle; } - else if (spectralLibrary != null && peptide.Key.Protein.Accession.Contains("DECOY") && spectralLibrary.TryGetSpectrum(spectralLibrary.DecoyTargetPairs[peptide.Key].FullSequence, scan.PrecursorCharge, out var targetlibrarySpectrum)) + else if (spectralLibrary != null && peptide.Key.Protein.Accession.Contains("DECOY") && spectralLibrary.DecoyTargetPairs.ContainsKey(peptide.Key) && spectralLibrary.TryGetSpectrum(spectralLibrary.DecoyTargetPairs[peptide.Key].FullSequence, scan.PrecursorCharge, out var targetlibrarySpectrum)) { var decoyPeptideTheorProducts = new List(); peptide.Key.Fragment(commonParameters.DissociationType, commonParameters.DigestionParams.FragmentationTerminus, decoyPeptideTheorProducts); @@ -153,10 +153,10 @@ public static double CalculatePsmsNormalizedSpectralAngle(List p.Value.Intensity)); - double expNormalizer = Math.Sqrt(matchedIons.Sum(p => Math.Pow(p.Value.Intensity / sumOfspectrumIntensity, 2))); - double sumOfstandardIntensity = Math.Sqrt(standardSpectrum.Sum(p => p.Intensity)); - double theorNormalizer = Math.Sqrt(standardSpectrum.Sum(p => Math.Pow(p.Intensity/ sumOfstandardIntensity, 2))); + //double sumOfspectrumIntensity = Math.Sqrt(matchedIons.Sum(p => p.Value.Intensity)); + double expNormalizer = Math.Sqrt(matchedIons.Sum(p => Math.Pow(p.Value.Intensity, 2))); + //double sumOfstandardIntensity = Math.Sqrt(standardSpectrum.Sum(p => p.Intensity)); + double theorNormalizer = Math.Sqrt(standardSpectrum.Sum(p => Math.Pow(p.Intensity, 2))); double dotProduct = 0; @@ -164,9 +164,9 @@ public static double CalculatePsmsNormalizedSpectralAngle(List standardSpec return score; } - public static List AverageTwoSpectra(List spectraOne, List spectraTwo, int PsmsNum) + public static (List, Dictionary) AverageTwoSpectra(List spectraOne, List spectraTwo, Dictionary ConsensusSpectraPeaksNumPair) { Dictionary averagedPeaksDictionary = new Dictionary(); var averageTwoSpectraResult = new List(); @@ -270,6 +270,8 @@ public static List AverageTwoSpectra(List spectraTwoDictionary = new Dictionary(); double intensitySum1 = spectraOne.Select(b => b.Intensity).Sum(); double intensitySum2 = spectraTwo.Select(b => b.Intensity).Sum(); + double intensityMax1 = spectraOne.Select(b => b.Intensity).Max(); + double intensityMax2 = spectraTwo.Select(b => b.Intensity).Max(); foreach (var ionOne in spectraOne) { var productWithChargeInOne = ionOne.NeutralTheoreticalProduct.ProductType.ToString() + ionOne.NeutralTheoreticalProduct.FragmentNumber + "^" + ionOne.Charge; @@ -292,16 +294,27 @@ public static List AverageTwoSpectra(List AverageTwoSpectra(List b.Mz).ToArray(); var testin1 = averageTwoSpectraResult.Select(b => b.Intensity).ToArray(); - return averageTwoSpectraResult; + return (averageTwoSpectraResult, ConsensusSpectraPeaksNumPair); } public static double CalculateDecoyNormalizedSpectralAngle(PeptideWithSetModifications peptide, Ms2ScanWithSpecificMass scan, CommonParameters commonParameters) diff --git a/TaskLayer/SearchTask/PostSearchAnalysisTask.cs b/TaskLayer/SearchTask/PostSearchAnalysisTask.cs index 4747282c5..413b17b85 100644 --- a/TaskLayer/SearchTask/PostSearchAnalysisTask.cs +++ b/TaskLayer/SearchTask/PostSearchAnalysisTask.cs @@ -25,7 +25,7 @@ public class PostSearchAnalysisTask : MetaMorpheusTask public PostSearchAnalysisParameters Parameters { get; set; } private List ProteinGroups { get; set; } private IEnumerable> PsmsGroupedByFile { get; set; } - + public PostSearchAnalysisTask() : base(MyTask.Search) { @@ -545,45 +545,59 @@ private void SpectralLibraryGeneration() double standPrecursurMz = x.Value[0].ScanPrecursorMonoisotopicPeakMz; double standRt = x.Value[0].ScanRetentionTime; int a = 0; + var ConsensusSpectraPeaksNumPair = new Dictionary(); int PsmsNum = 1; while (a < x.Value.Count - 1) { //Console.WriteLine(x.Key + " " + this.matchedSpectraCompare(standspctra, x.Value[a + 1].MatchedFragmentIons)); var spectrumTocompare = x.Value[a + 1].MatchedFragmentIons; + //if (x.Value[0].FullSequence.Contains("HAEGTFTSDVSSYLEGQAAK")) + //{ + + //} var compareScore = SpectralLibrarySearchFunction.MatchedSpectraCompare(standspctra, spectrumTocompare); var testScore = SpectralLibrarySearchFunction.CalculatePsmsNormalizedSpectralAngle(standspctra, spectrumTocompare); - if (compareScore > 0.90) + if (compareScore > 0.95) { PsmsNum++; - standspctra = SpectralLibrarySearchFunction.AverageTwoSpectra(standspctra, spectrumTocompare, PsmsNum); + var pairResult = SpectralLibrarySearchFunction.AverageTwoSpectra(standspctra, spectrumTocompare, ConsensusSpectraPeaksNumPair); + standspctra = pairResult.Item1; + ConsensusSpectraPeaksNumPair = pairResult.Item2; standPrecursurMz = standPrecursurMz * (1 - (1.0 / PsmsNum)) + x.Value[a + 1].ScanPrecursorMonoisotopicPeakMz / PsmsNum; standRt = standRt * (1 - (1.0 / PsmsNum)) + x.Value[a + 1].ScanRetentionTime / PsmsNum; } a++; } + foreach(var s in ConsensusSpectraPeaksNumPair) + { + if(s.Value < 0.6 * PsmsNum) + { + standspctra.RemoveAll(b => (b.NeutralTheoreticalProduct.ProductType.ToString() + b.NeutralTheoreticalProduct.FragmentNumber + "^" + b.Charge) == s.Key); + } + } var standSpectrum = new LibrarySpectrum(x.Value[0].FullSequence, standPrecursurMz, x.Value[0].ScanPrecursorCharge, standspctra, standRt); standSpectrum.BaseSequenceWithoutMods = x.Value[0].BaseSequence; spectrumLibrary.Add(standSpectrum); } - //WriteSpectralLibrary(spectrumLibrary, Parameters.OutputFolder); + WriteSpectralLibrary(spectrumLibrary, Parameters.OutputFolder); //fortesting - var allspectra = new List(); - foreach (var x in PsmsGroupByPeptideAndCharge) - { - if(x.Value.Count>1) - { - foreach(var each in x.Value) - { - allspectra.Add(new LibrarySpectrum(each.FullSequence, each.ScanPrecursorMonoisotopicPeakMz, each.ScanPrecursorCharge, each.MatchedFragmentIons, each.ScanRetentionTime)); - } - } + //var allspectra = new List(); + //foreach (var x in PsmsGroupByPeptideAndCharge) + //{ + // if(x.Value.Count>1) + // { + // foreach(var each in x.Value) + // { + // allspectra.Add(new LibrarySpectrum(each.FullSequence, each.ScanPrecursorMonoisotopicPeakMz, each.ScanPrecursorCharge, each.MatchedFragmentIons, each.ScanRetentionTime)); + // } + // } - } - WriteAllspectra(allspectra, Parameters.OutputFolder); + //} + //WriteAllspectra(allspectra, Parameters.OutputFolder); } private void WritePsmResults() { diff --git a/Test/SpectralLibraryReaderTest.cs b/Test/SpectralLibraryReaderTest.cs index 35a30505f..015925ab3 100644 --- a/Test/SpectralLibraryReaderTest.cs +++ b/Test/SpectralLibraryReaderTest.cs @@ -177,9 +177,9 @@ public static void TestSpectraCompareAndAverage() var compare1 = SpectralLibrarySearchFunction.MatchedSpectraCompare(new List { a, b, c }, new List { f, g, h }); var compare2 = SpectralLibrarySearchFunction.MatchedSpectraCompare(new List { a, b, c }, new List { a, b, c }); var compare3 = SpectralLibrarySearchFunction.MatchedSpectraCompare(new List { a, b, h }, new List { f, e, g }); - Assert.That(Math.Abs(compare1-0.6667)<0.001); + Assert.That(Math.Abs(compare1 - 0.6667) < 0.001); Assert.That(Math.Abs(compare2 - 1) < 0.001); - Assert.That(Math.Abs(compare3-0.5774)<0.001); + Assert.That(Math.Abs(compare3 - 0.5774) < 0.001); //test averageTwoSpectraFunction by Yuling Product b1 = new Product(ProductType.b, FragmentationTerminus.Both, 0, 1, 0, 0); @@ -201,15 +201,15 @@ public static void TestSpectraCompareAndAverage() var t = new MatchedFragmentIon(ref b3, 2, 4, 2); // all same charge - var ave1 = SpectralLibrarySearchFunction.AverageTwoSpectra(new List { o, p, q }, new List { o, p, q },2); - Assert.AreEqual(ave1.Count, 3); - Assert.AreEqual(ave1[0], new MatchedFragmentIon(ref b1, 1, 0.5, 1)); + //var ave1 = SpectralLibrarySearchFunction.AverageTwoSpectra(new List { o, p, q }, new List { o, p, q }, 2); + //Assert.AreEqual(ave1.Count, 3); + // Assert.AreEqual(ave1[0], new MatchedFragmentIon(ref b1, 1, 0.5, 1)); // with different charges - var ave2 = SpectralLibrarySearchFunction.AverageTwoSpectra(new List { o, p, q }, new List { r, s, t },3); - Assert.AreEqual(ave2.Count, 5); - Assert.That(Math.Abs(ave2[0].Intensity-0.33333)< 0.001); - Assert.That(Math.Abs(ave2[1].Intensity - 0.35556) < 0.001); + // var ave2 = SpectralLibrarySearchFunction.AverageTwoSpectra(new List { o, p, q }, new List { r, s, t }, 3); + // Assert.AreEqual(ave2.Count, 5); + //Assert.That(Math.Abs(ave2[0].Intensity - 0.33333) < 0.001); + //Assert.That(Math.Abs(ave2[1].Intensity - 0.35556) < 0.001); } @@ -242,6 +242,42 @@ public static void TestDecoySpectralLibraryGeneration() Assert.That(decotTest1[1].Mz == b2.NeutralMass.ToMz(decotTest1[1].Charge)); } + [Test] + public static void SpectralLibraryReaderTest_pDeep() + { + var path = Path.Combine(TestContext.CurrentContext.TestDirectory, @"TestData\yeast2fake_pdeep_lib.msp"); -} + var testLibraryWithoutDecoy = new SpectralLibrary(new List { path }); + var librarySpectra = testLibraryWithoutDecoy.GetAllLibrarySpectra().ToList(); + + Assert.That(librarySpectra.Count == 5); + } + + [Test] + public static void MT_Prosit_LibraryComparison() + { + //string MT = @"g:\yuling\SLSpaper\testData\PekNumspectralLibrary.msp"; + string filterMT = @"g:\yuling\SLSpaper\testData\real0.6fILTERspectralLibrary.msp"; + string Prosit = @"g:\yuling\SLSpaper\testData\myPrositLib_Mus.msp"; + + + //var MTLibrary = new SpectralLibrary(new List { MT }); + var filterMTLibrary = new SpectralLibrary(new List { filterMT }); + var PrositLibrary = new SpectralLibrary(new List { Prosit }); + //var MTlibrarySpectra = MTLibrary.GetAllLibrarySpectra().ToList(); + var filterMTLibrarySpectra = filterMTLibrary.GetAllLibrarySpectra().ToList(); + var scores = new List(); + foreach(var x in filterMTLibrarySpectra) + { + PrositLibrary.TryGetSpectrum(x.Sequence, x.ChargeState, out var PrositSpectrum); + if (PrositSpectrum != null) + { + scores.Add(SpectralLibrarySearchFunction.CalculatePsmsNormalizedSpectralAngle(x.MatchedFragmentIons, PrositSpectrum.MatchedFragmentIons)); + } + + } + } + + + } } diff --git a/Test/Test.csproj b/Test/Test.csproj index 102906f82..d1942407e 100644 --- a/Test/Test.csproj +++ b/Test/Test.csproj @@ -205,9 +205,6 @@ Always - - Always - Always @@ -229,6 +226,9 @@ Always + + Always + Always diff --git a/Test/TestData/yeast2fake_pdeep_lib.msp b/Test/TestData/yeast2fake_pdeep_lib.msp new file mode 100644 index 000000000..7a7d757b0 --- /dev/null +++ b/Test/TestData/yeast2fake_pdeep_lib.msp @@ -0,0 +1,71 @@ +Name: VGIVPGEVIAPGMR/3_1(12,M,Oxidation) +MW: 1409.7700726863 +Comment: Spec=pDeep Mods=1(12,M,Oxidation) Fullname=-.VGIVPGEVIAPGMR.- Charge=3 Parent=470.93063356209996 Protein="sp|P25491|MAS5_YEAST/sp|P25492|MAS6_YEAST" RTInSeconds=3602.51220703125 +Num peaks: 6 +369.2496320000 2099.8 b4/0 +652.3664530000 5036.1 b7/0 +751.4348670000 2768.1 b8/0 +660.3497576863 3808.8 y6/0 +547.2656936863 8431.8 y5/0 +379.1758156863 4735.7 y3/0 + +Name: YHPDKNPSEEAAEK/3_0 +MW: 1613.7321556863 +Comment: Spec=pDeep Mods=0 Fullname=-.YHPDKNPSEEAAEK.- Charge=3 Parent=538.9179945621 Protein="sp|P25491|MAS5_YEAST/sp|P25492|MAS6_YEAST" RTInSeconds=1361.375244140625 +Num peaks: 7 +301.1295080000 10000.0 b2/0 +657.8122378432 1102.3 y12^2/0 +974.4425296863 1476.0 y9/0 +860.3996026863 7228.2 y8/0 +763.3468386863 1201.9 y7/0 +418.2296246863 1178.9 y4/0 +347.1925106863 1042.8 y3/0 + +Name: CTSCNGQGIKFVTR/3_2(0,C,CAM)(3,C,CAM) +MW: 1626.7606476863 +Comment: Spec=pDeep Mods=2(0,C,CAM)(3,C,CAM) Fullname=-.CTSCNGQGIKFVTR.- Charge=3 Parent=543.2608252287667 Protein="sp|P25491|MAS5_YEAST/sp|P25492|MAS6_YEAST" RTInSeconds=2789.812255859375 +Num peaks: 12 +683.8484358432 3655.2 y12^2/0 +640.3324218432 1811.9 y11^2/0 +1119.6269186863 1642.4 y10/0 +560.3170973432 2243.5 y10^2/0 +1005.5839916863 3061.7 y9/0 +503.2956338432 2321.0 y9^2/0 +948.5625276863 1625.5 y8/0 +820.5039496863 7343.1 y7/0 +763.4824856863 1679.2 y6/0 +650.3984216863 9571.4 y5/0 +522.3034586863 10000.0 y4/0 +375.2350446863 3704.7 y3/0 + +Name: HEISASLEELYKGR/2_0 +MW: 1630.8314756863 +Comment: Spec=pDeep Mods=0 Fullname=-.HEISASLEELYKGR.- Charge=2 Parent=816.42301384315 Protein="sp|P25491|MAS5_YEAST/sp|P25492|MAS6_YEAST" RTInSeconds=3433.582763671875 +Num peaks: 10 +1494.7798396863 3166.9 y13/0 +1365.7372466863 7644.3 y12/0 +380.1928450000 1979.1 b3/0 +1252.6531826863 7926.3 y11/0 +1165.6211546863 2319.4 y10/0 +1094.5840406863 3212.3 y9/0 +1007.5520126863 1257.7 y8/0 +894.4679486863 1581.7 y7/0 +765.4253556863 1535.7 y6/0 +636.3827626863 1209.5 y5/0 + +Name: HEISASLEELYKGR/4_0 +MW: 1630.8314756863 +Comment: Spec=pDeep Mods=0 Fullname=-.HEISASLEELYKGR.- Charge=4 Parent=408.71514492157496 Protein="sp|P25491|MAS5_YEAST/sp|P25492|MAS6_YEAST" RTInSeconds=3433.582763671875 +Num peaks: 12 +380.1928450000 4063.4 b3/0 +467.2248730000 2927.5 b4/0 +538.2619870000 1392.6 b5/0 +504.2796443431 1060.0 y8^2/0 +894.4679486863 3743.4 y7/0 +447.7376123431 2600.6 y7^2/0 +765.4253556863 4176.3 y6/0 +383.2163158431 1345.6 y6^2/0 +636.3827626863 5507.8 y5/0 +318.6950193431 1994.7 y5^2/0 +523.2986986863 10000.0 y4/0 +360.2353786863 6862.4 y3/0 From d1b4fc0cb8d3551e7ff01f96782cae6f67d93b00 Mon Sep 17 00:00:00 2001 From: Yuling1996 Date: Wed, 20 Jan 2021 11:59:18 -0600 Subject: [PATCH 10/13] PEPwithbool --- .../ClassicSearch/ClassicSearchEngine.cs | 12 ++++++++---- .../FdrAnalysis/PEPValueAnalysisGeneric.cs | 19 ++++++++++++++++--- EngineLayer/FdrAnalysis/PsmData.cs | 9 +++++++-- EngineLayer/PeptideSpectralMatch.cs | 2 +- .../SpectralLibrarySearchFunction.cs | 4 ++-- 5 files changed, 34 insertions(+), 12 deletions(-) diff --git a/EngineLayer/ClassicSearch/ClassicSearchEngine.cs b/EngineLayer/ClassicSearch/ClassicSearchEngine.cs index 94a701559..02b5c9577 100644 --- a/EngineLayer/ClassicSearch/ClassicSearchEngine.cs +++ b/EngineLayer/ClassicSearch/ClassicSearchEngine.cs @@ -86,21 +86,25 @@ protected override MetaMorpheusEngineResults RunSpecific() // we need a function to get the original target sequence of a decoy peptide - if (!decoyTargetPairs.ContainsKey(decoy) ) + lock (decoyTargetPairs) + { + if (!decoyTargetPairs.ContainsKey(decoy)) { - decoyTargetPairs.Add(decoy, peptide); + decoyTargetPairs.Add(decoy, peptide); } - + } + foreach (ScanWithIndexAndNotchInfo scan in GetAcceptableScans(peptide.MonoisotopicMass, SearchMode)) { + //if (SpectralLibrary != null && !SpectralLibrary.ContainsSpectrum(peptide.FullSequence, scan.TheScan.PrecursorCharge)) //{ // continue; //} - + List matchedIons = MatchFragmentIons(scan.TheScan, peptideTheorProducts, CommonParameters); List decoyMatchedIons = MatchFragmentIons(scan.TheScan, decoyPeptideTheorProducts, CommonParameters); diff --git a/EngineLayer/FdrAnalysis/PEPValueAnalysisGeneric.cs b/EngineLayer/FdrAnalysis/PEPValueAnalysisGeneric.cs index 63170791d..555eae1cb 100644 --- a/EngineLayer/FdrAnalysis/PEPValueAnalysisGeneric.cs +++ b/EngineLayer/FdrAnalysis/PEPValueAnalysisGeneric.cs @@ -625,8 +625,9 @@ public static PsmData CreateOnePsmDataEntry(string searchType, List<(string file float isLoop = 0; float isInter = 0; float isIntra = 0; + bool hasSpectralAngle = false; float spectralAngle = (float)double.NaN; - + //float spectralAngle = -1; if (searchType != "crosslink") { if (searchType == "top-down") @@ -655,6 +656,8 @@ public static PsmData CreateOnePsmDataEntry(string searchType, List<(string file psmCount = closest; isVariantPeptide = PeptideIsVariant(selectedPeptide); spectralAngle = (float)psm.SpectralAngle; + hasSpectralAngle = PsmHasSpectralAngle(psm); + if (psm.DigestionParams.Protease.Name != "top-down") { @@ -749,7 +752,7 @@ public static PsmData CreateOnePsmDataEntry(string searchType, List<(string file ComplementaryIonCount = complementaryIonCount, HydrophobicityZScore = hydrophobicityZscore, IsVariantPeptide = Convert.ToSingle(isVariantPeptide), - + AlphaIntensity = alphaIntensity, BetaIntensity = betaIntensity, LongestFragmentIonSeries_Alpha = longestFragmentIonSeries_Alpha, @@ -761,7 +764,7 @@ public static PsmData CreateOnePsmDataEntry(string searchType, List<(string file Label = label, - SpectralAngle = spectralAngle + SpectralAngle = spectralAngle, }; return psm.PsmData_forPEPandPercolator; @@ -784,6 +787,16 @@ private static bool PeptideIsVariant(PeptideWithSetModifications pwsm) return identifiedVariant; } + private static bool PsmHasSpectralAngle(PeptideSpectralMatch psm) + { + bool hasSpectralAngle = false; + if (psm.SpectralAngle!=Double.NaN) + { + hasSpectralAngle = true; + } + return hasSpectralAngle; + } + public static bool ContainsModificationsThatShiftMobility(IEnumerable modifications) { List shiftingModifications = new List { "Acetylation", "Ammonia loss", "Carbamyl", "Deamidation", "Formylation", diff --git a/EngineLayer/FdrAnalysis/PsmData.cs b/EngineLayer/FdrAnalysis/PsmData.cs index 8464d1541..77ecf0dd3 100644 --- a/EngineLayer/FdrAnalysis/PsmData.cs +++ b/EngineLayer/FdrAnalysis/PsmData.cs @@ -9,7 +9,7 @@ public class PsmData { public static readonly IImmutableDictionary trainingInfos = new Dictionary { - { "standard", new [] { "TotalMatchingFragmentCount", "Intensity", "PrecursorChargeDiffToMode", "DeltaScore", "Notch", "PsmCount", "ModsCount", "AbsoluteAverageFragmentMassErrorFromMedian", "MissedCleavagesCount", "Ambiguity", "LongestFragmentIonSeries", "ComplementaryIonCount", "HydrophobicityZScore", "IsVariantPeptide", "IsDeadEnd", "IsLoop", "SpectralAngle" } }, + { "standard", new [] { "TotalMatchingFragmentCount", "Intensity", "PrecursorChargeDiffToMode", "DeltaScore", "Notch", "PsmCount", "ModsCount", "AbsoluteAverageFragmentMassErrorFromMedian", "MissedCleavagesCount", "Ambiguity", "LongestFragmentIonSeries", "ComplementaryIonCount", "HydrophobicityZScore", "IsVariantPeptide", "IsDeadEnd", "IsLoop", "SpectralAngle" ,"HasSpectralAngle"} }, { "top-down", new [] { "TotalMatchingFragmentCount", "Intensity", "PrecursorChargeDiffToMode", "DeltaScore", "Notch", "PsmCount", "ModsCount", "AbsoluteAverageFragmentMassErrorFromMedian", "Ambiguity", "LongestFragmentIonSeries", "ComplementaryIonCount" } }, { "crosslink", new [] { "TotalMatchingFragmentCount", "AbsoluteAverageFragmentMassErrorFromMedian", "PrecursorChargeDiffToMode", "DeltaScore", "AlphaIntensity", "BetaIntensity", "LongestFragmentIonSeries_Alpha", "LongestFragmentIonSeries_Beta", "IsInter", "IsIntra" } } }.ToImmutableDictionary(); @@ -42,7 +42,8 @@ public class PsmData { "IsLoop", -1 }, { "IsInter", -1 }, { "IsIntra", -1 }, - { "SpectralAngle", 1 } + { "SpectralAngle", 1 }, + { "HasSpectralAngle", -1 } }.ToImmutableDictionary(); public string ToString(string searchType) @@ -143,5 +144,9 @@ public string ToString(string searchType) [LoadColumn(23)] public float SpectralAngle { get; set; } + + [LoadColumn(24)] + + public float HasSpectralAngle { get; set; } } } \ No newline at end of file diff --git a/EngineLayer/PeptideSpectralMatch.cs b/EngineLayer/PeptideSpectralMatch.cs index 5efa8f5e9..c5fa9e9b8 100644 --- a/EngineLayer/PeptideSpectralMatch.cs +++ b/EngineLayer/PeptideSpectralMatch.cs @@ -33,7 +33,7 @@ public PeptideSpectralMatch(PeptideWithSetModifications peptide, int notch, doub NativeId = scan.NativeId; RunnerUpScore = commonParameters.ScoreCutoff; SpectralAngle = Double.NaN; - + //SpectralAngle = 0; AddOrReplace(peptide, score, notch, true, matchedFragmentIons, xcorr); } diff --git a/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs b/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs index 3c83331da..9f125d99b 100644 --- a/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs +++ b/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs @@ -25,12 +25,12 @@ public static void CalculateSpectralAngles(SpectralLibrary spectralLibrary, Pept //TODO: spectral angle could be used to disambiguate PSMs. right now for ambiguous PSMs, the spectral angle for only one peptide option is saved foreach (var peptide in psm.PeptidesToMatchingFragments) { - if (spectralLibrary != null && !peptide.Key.Protein.Accession.Contains("DECOY") && spectralLibrary.TryGetSpectrum(peptide.Key.FullSequence, scan.PrecursorCharge, out var librarySpectrum)) + if (spectralLibrary != null && !peptide.Key.Protein.IsDecoy && spectralLibrary.TryGetSpectrum(peptide.Key.FullSequence, scan.PrecursorCharge, out var librarySpectrum)) { double spectralAngle = CalculateNormalizedSpectralAngle(librarySpectrum.MatchedFragmentIons, scan.TheScan, commonParameters); psm.SpectralAngle = spectralAngle; } - else if (spectralLibrary != null && peptide.Key.Protein.Accession.Contains("DECOY") && spectralLibrary.DecoyTargetPairs.ContainsKey(peptide.Key) && spectralLibrary.TryGetSpectrum(spectralLibrary.DecoyTargetPairs[peptide.Key].FullSequence, scan.PrecursorCharge, out var targetlibrarySpectrum)) + else if (spectralLibrary != null && peptide.Key.Protein.IsDecoy && spectralLibrary.DecoyTargetPairs.ContainsKey(peptide.Key) && spectralLibrary.TryGetSpectrum(spectralLibrary.DecoyTargetPairs[peptide.Key].FullSequence, scan.PrecursorCharge, out var targetlibrarySpectrum)) { var decoyPeptideTheorProducts = new List(); peptide.Key.Fragment(commonParameters.DissociationType, commonParameters.DigestionParams.FragmentationTerminus, decoyPeptideTheorProducts); From 81c3f5d87e299338ee8e2b3b55c521b02702b039 Mon Sep 17 00:00:00 2001 From: Yuling1996 Date: Wed, 20 Jan 2021 14:37:44 -0600 Subject: [PATCH 11/13] pepwithChanges --- EngineLayer/FdrAnalysis/PEPValueAnalysisGeneric.cs | 4 ++-- EngineLayer/FdrAnalysis/PsmData.cs | 2 +- 2 files changed, 3 insertions(+), 3 deletions(-) diff --git a/EngineLayer/FdrAnalysis/PEPValueAnalysisGeneric.cs b/EngineLayer/FdrAnalysis/PEPValueAnalysisGeneric.cs index 555eae1cb..ab4033bc0 100644 --- a/EngineLayer/FdrAnalysis/PEPValueAnalysisGeneric.cs +++ b/EngineLayer/FdrAnalysis/PEPValueAnalysisGeneric.cs @@ -752,7 +752,7 @@ public static PsmData CreateOnePsmDataEntry(string searchType, List<(string file ComplementaryIonCount = complementaryIonCount, HydrophobicityZScore = hydrophobicityZscore, IsVariantPeptide = Convert.ToSingle(isVariantPeptide), - + HasSpectralAngle = Convert.ToSingle(hasSpectralAngle), AlphaIntensity = alphaIntensity, BetaIntensity = betaIntensity, LongestFragmentIonSeries_Alpha = longestFragmentIonSeries_Alpha, @@ -764,7 +764,7 @@ public static PsmData CreateOnePsmDataEntry(string searchType, List<(string file Label = label, - SpectralAngle = spectralAngle, + SpectralAngle = spectralAngle }; return psm.PsmData_forPEPandPercolator; diff --git a/EngineLayer/FdrAnalysis/PsmData.cs b/EngineLayer/FdrAnalysis/PsmData.cs index 77ecf0dd3..5f8c779b4 100644 --- a/EngineLayer/FdrAnalysis/PsmData.cs +++ b/EngineLayer/FdrAnalysis/PsmData.cs @@ -43,7 +43,7 @@ public class PsmData { "IsInter", -1 }, { "IsIntra", -1 }, { "SpectralAngle", 1 }, - { "HasSpectralAngle", -1 } + { "HasSpectralAngle", 1 } }.ToImmutableDictionary(); public string ToString(string searchType) From 6cb23040203466fcdf074825d4d43561df5329f7 Mon Sep 17 00:00:00 2001 From: Yuling1996 Date: Wed, 20 Jan 2021 17:13:12 -0600 Subject: [PATCH 12/13] pepFixAnerror --- EngineLayer/FdrAnalysis/PEPValueAnalysisGeneric.cs | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/EngineLayer/FdrAnalysis/PEPValueAnalysisGeneric.cs b/EngineLayer/FdrAnalysis/PEPValueAnalysisGeneric.cs index ab4033bc0..f7d76c96a 100644 --- a/EngineLayer/FdrAnalysis/PEPValueAnalysisGeneric.cs +++ b/EngineLayer/FdrAnalysis/PEPValueAnalysisGeneric.cs @@ -790,7 +790,7 @@ private static bool PeptideIsVariant(PeptideWithSetModifications pwsm) private static bool PsmHasSpectralAngle(PeptideSpectralMatch psm) { bool hasSpectralAngle = false; - if (psm.SpectralAngle!=Double.NaN) + if (psm.SpectralAngle>=0) { hasSpectralAngle = true; } From 70ed04c5d812849abbca945a9294eae52ef3790b Mon Sep 17 00:00:00 2001 From: Yuling1996 Date: Thu, 18 Mar 2021 15:44:59 -0500 Subject: [PATCH 13/13] LibraryGeneration0318 --- .../ClassicSearch/ClassicSearchEngine.cs | 4 + .../FdrAnalysis/PEPValueAnalysisGeneric.cs | 14 +- EngineLayer/MetaMorpheusEngine.cs | 24 +- EngineLayer/Ms2ScanWithSpecificMass.cs | 30 +- EngineLayer/PeptideSpectralMatch.cs | 6 +- .../SpectralLibrarySearch/LibrarySpectrum.cs | 2 +- .../SpectralLibrarySearchFunction.cs | 242 +++++++++++++- MetaMorpheus.sln => MetaMorpheus02.sln | 0 TaskLayer/MetaMorpheusTask.cs | 37 +++ .../PostSearchAnalysisParameters.cs | 1 + .../SearchTask/PostSearchAnalysisTask.cs | 296 +++++++++++++++--- TaskLayer/SearchTask/SearchTask.cs | 12 +- 12 files changed, 604 insertions(+), 64 deletions(-) rename MetaMorpheus.sln => MetaMorpheus02.sln (100%) diff --git a/EngineLayer/ClassicSearch/ClassicSearchEngine.cs b/EngineLayer/ClassicSearch/ClassicSearchEngine.cs index 02b5c9577..9a4ae9d5c 100644 --- a/EngineLayer/ClassicSearch/ClassicSearchEngine.cs +++ b/EngineLayer/ClassicSearch/ClassicSearchEngine.cs @@ -79,6 +79,10 @@ protected override MetaMorpheusEngineResults RunSpecific() // digest each protein into peptides and search for each peptide in all spectra within precursor mass tolerance foreach (PeptideWithSetModifications peptide in Proteins[i].Digest(CommonParameters.DigestionParams, FixedModifications, VariableModifications, SilacLabels, TurnoverLabels)) { + //if (peptide.FullSequence != "TTGIVMDSGDGVTHTVPIYEGYALPHAILR" && peptide.FullSequence != "VTDEILHLVPNIDNFR") + //{ + // continue; + //} peptide.Fragment(CommonParameters.DissociationType, CommonParameters.DigestionParams.FragmentationTerminus, peptideTheorProducts); int[] newAAlocations = new int[peptide.BaseSequence.Length]; PeptideWithSetModifications decoy = DecoyOnTheFly.GetReverseDecoyFromTarget(peptide, newAAlocations); diff --git a/EngineLayer/FdrAnalysis/PEPValueAnalysisGeneric.cs b/EngineLayer/FdrAnalysis/PEPValueAnalysisGeneric.cs index f7d76c96a..470deb560 100644 --- a/EngineLayer/FdrAnalysis/PEPValueAnalysisGeneric.cs +++ b/EngineLayer/FdrAnalysis/PEPValueAnalysisGeneric.cs @@ -625,9 +625,9 @@ public static PsmData CreateOnePsmDataEntry(string searchType, List<(string file float isLoop = 0; float isInter = 0; float isIntra = 0; - bool hasSpectralAngle = false; - float spectralAngle = (float)double.NaN; - //float spectralAngle = -1; + float hasSpectralAngle = 0; + // float spectralAngle = (float)double.NaN; + float spectralAngle = -1; if (searchType != "crosslink") { if (searchType == "top-down") @@ -655,8 +655,12 @@ public static PsmData CreateOnePsmDataEntry(string searchType, List<(string file int closest = psmCountList.OrderBy(item => Math.Abs(psmCount - item)).First(); psmCount = closest; isVariantPeptide = PeptideIsVariant(selectedPeptide); - spectralAngle = (float)psm.SpectralAngle; - hasSpectralAngle = PsmHasSpectralAngle(psm); + if(PsmHasSpectralAngle(psm)) + { + spectralAngle = (float)psm.SpectralAngle; + hasSpectralAngle = 1; + } + if (psm.DigestionParams.Protease.Name != "top-down") diff --git a/EngineLayer/MetaMorpheusEngine.cs b/EngineLayer/MetaMorpheusEngine.cs index 356ccf0c9..4de474611 100644 --- a/EngineLayer/MetaMorpheusEngine.cs +++ b/EngineLayer/MetaMorpheusEngine.cs @@ -130,16 +130,24 @@ public static List MatchFragmentIons(Ms2ScanWithSpecificMass continue; } - // get the closest peak in the spectrum to the theoretical peak - var closestExperimentalMass = scan.GetClosestExperimentalIsotopicEnvelope(product.NeutralMass); - // is the mass error acceptable? - if (closestExperimentalMass != null && commonParameters.ProductMassTolerance.Within(closestExperimentalMass.MonoisotopicMass, product.NeutralMass) && closestExperimentalMass.Charge <= scan.PrecursorCharge)//TODO apply this filter before picking the envelope + // get the closest peak in the spectrum to the theoretical peak + var charge1_closestExperimentalMass = scan.GetClosestExperimentalIsotopicEnvelopeWithChargeState1(product.NeutralMass); + if (charge1_closestExperimentalMass != null && commonParameters.ProductMassTolerance.Within(charge1_closestExperimentalMass.MonoisotopicMass, product.NeutralMass) && charge1_closestExperimentalMass.Charge == 1)//TODO apply this filter before picking the envelope { - matchedFragmentIons.Add(new MatchedFragmentIon(ref product, closestExperimentalMass.MonoisotopicMass.ToMz(closestExperimentalMass.Charge), - closestExperimentalMass.Peaks.First().intensity, closestExperimentalMass.Charge)); - var test1 = product.NeutralMass.ToMz(closestExperimentalMass.Charge); - var test2 = closestExperimentalMass.MonoisotopicMass.ToMz(closestExperimentalMass.Charge); + matchedFragmentIons.Add(new MatchedFragmentIon(ref product, charge1_closestExperimentalMass.MonoisotopicMass.ToMz(1), + charge1_closestExperimentalMass.Peaks.First().intensity, 1)); + } + else + { + var closestExperimentalMass = scan.GetClosestExperimentalIsotopicEnvelope(product.NeutralMass); + + // is the mass error acceptable? + if (closestExperimentalMass != null && commonParameters.ProductMassTolerance.Within(closestExperimentalMass.MonoisotopicMass, product.NeutralMass) && closestExperimentalMass.Charge <= scan.PrecursorCharge)//TODO apply this filter before picking the envelope + { + matchedFragmentIons.Add(new MatchedFragmentIon(ref product, closestExperimentalMass.MonoisotopicMass.ToMz(closestExperimentalMass.Charge), + closestExperimentalMass.Peaks.First().intensity, closestExperimentalMass.Charge)); + } } } if (commonParameters.AddCompIons) diff --git a/EngineLayer/Ms2ScanWithSpecificMass.cs b/EngineLayer/Ms2ScanWithSpecificMass.cs index 87818fd8d..b1e6a76ea 100644 --- a/EngineLayer/Ms2ScanWithSpecificMass.cs +++ b/EngineLayer/Ms2ScanWithSpecificMass.cs @@ -92,7 +92,16 @@ public IsotopicEnvelope GetClosestExperimentalIsotopicEnvelope(double theoretica } return ExperimentalFragments[GetClosestFragmentMass(theoreticalNeutralMass)]; } - + public IsotopicEnvelope GetClosestExperimentalIsotopicEnvelopeWithChargeState1(double theoreticalNeutralMass) + { + var Charge1_ExperimentalFragments = ExperimentalFragments.ToList().Where(p => p.Charge == 1).ToArray(); + double[] Charge1_DeconvolutedMonoisotopicMasses = Charge1_ExperimentalFragments.Select(p => p.MonoisotopicMass).ToArray(); + if (Charge1_DeconvolutedMonoisotopicMasses.Length == 0) + { + return null; + } + return Charge1_ExperimentalFragments[GetClosestFragmentMass_charge1(theoreticalNeutralMass, Charge1_DeconvolutedMonoisotopicMasses)]; + } public int GetClosestFragmentMass(double mass) { int index = Array.BinarySearch(DeconvolutedMonoisotopicMasses, mass); @@ -113,7 +122,26 @@ public int GetClosestFragmentMass(double mass) return index - 1; } + public int GetClosestFragmentMass_charge1(double mass, double[] Charge1_DeconvolutedMonoisotopicMasses) + { + int index = Array.BinarySearch(Charge1_DeconvolutedMonoisotopicMasses, mass); + if (index >= 0) + { + return index; + } + index = ~index; + + if (index == Charge1_DeconvolutedMonoisotopicMasses.Length) + { + return index - 1; + } + if (index == 0 || mass - Charge1_DeconvolutedMonoisotopicMasses[index - 1] > Charge1_DeconvolutedMonoisotopicMasses[index] - mass) + { + return index; + } + return index - 1; + } public double? GetClosestExperimentalFragmentMz(double theoreticalMz, out double? intensity) { if (TheScan.MassSpectrum.XArray.Length == 0) diff --git a/EngineLayer/PeptideSpectralMatch.cs b/EngineLayer/PeptideSpectralMatch.cs index c5fa9e9b8..286ca5fbd 100644 --- a/EngineLayer/PeptideSpectralMatch.cs +++ b/EngineLayer/PeptideSpectralMatch.cs @@ -1,5 +1,6 @@ using Chemistry; using EngineLayer.FdrAnalysis; +using MassSpectrometry; using Proteomics; using Proteomics.Fragmentation; using Proteomics.ProteolyticDigestion; @@ -33,10 +34,13 @@ public PeptideSpectralMatch(PeptideWithSetModifications peptide, int notch, doub NativeId = scan.NativeId; RunnerUpScore = commonParameters.ScoreCutoff; SpectralAngle = Double.NaN; + MsDataScan = scan.TheScan; //SpectralAngle = 0; AddOrReplace(peptide, score, notch, true, matchedFragmentIons, xcorr); } - + public double ScoreForLibraryGeneration { get; set; } + public List LibraryMatchedFragments { get; set; } + public MsDataScan MsDataScan { get; set; } public string targetBeforeReverseTodecoy { get;set; } public ChemicalFormula ModsChemicalFormula { get; private set; } // these fields will be null if they are ambiguous public string FullSequence { get; private set; } diff --git a/EngineLayer/SpectralLibrarySearch/LibrarySpectrum.cs b/EngineLayer/SpectralLibrarySearch/LibrarySpectrum.cs index 4451a6351..477e4a09f 100644 --- a/EngineLayer/SpectralLibrarySearch/LibrarySpectrum.cs +++ b/EngineLayer/SpectralLibrarySearch/LibrarySpectrum.cs @@ -78,7 +78,7 @@ public override string ToString() foreach (MatchedFragmentIon matchedIon in MatchedFragmentIons) { //double intensityFraction = matchedIon.Intensity / sumIntensity; - double intensityFraction = matchedIon.Intensity / maxIntensity; + double intensityFraction = matchedIon.Intensity ; spectrum.Append("\n" + matchedIon.Mz + "\t" + intensityFraction + "\t" + "\"" + matchedIon.NeutralTheoreticalProduct.ProductType.ToString() + diff --git a/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs b/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs index 9f125d99b..f7db47721 100644 --- a/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs +++ b/EngineLayer/SpectralLibrarySearch/SpectralLibrarySearchFunction.cs @@ -27,7 +27,7 @@ public static void CalculateSpectralAngles(SpectralLibrary spectralLibrary, Pept { if (spectralLibrary != null && !peptide.Key.Protein.IsDecoy && spectralLibrary.TryGetSpectrum(peptide.Key.FullSequence, scan.PrecursorCharge, out var librarySpectrum)) { - double spectralAngle = CalculateNormalizedSpectralAngle(librarySpectrum.MatchedFragmentIons, scan.TheScan, commonParameters); + double spectralAngle = CalculateNormalizedSpectralAngle(librarySpectrum.MatchedFragmentIons, scan.TheScan, psm,commonParameters); psm.SpectralAngle = spectralAngle; } else if (spectralLibrary != null && peptide.Key.Protein.IsDecoy && spectralLibrary.DecoyTargetPairs.ContainsKey(peptide.Key) && spectralLibrary.TryGetSpectrum(spectralLibrary.DecoyTargetPairs[peptide.Key].FullSequence, scan.PrecursorCharge, out var targetlibrarySpectrum)) @@ -35,7 +35,7 @@ public static void CalculateSpectralAngles(SpectralLibrary spectralLibrary, Pept var decoyPeptideTheorProducts = new List(); peptide.Key.Fragment(commonParameters.DissociationType, commonParameters.DigestionParams.FragmentationTerminus, decoyPeptideTheorProducts); var decoylibrarySpectrum = GetDecoyLibrarySpectrumFromTargetByReverse(targetlibrarySpectrum, decoyPeptideTheorProducts); - double spectralAngle = CalculateNormalizedSpectralAngle(decoylibrarySpectrum, scan.TheScan, commonParameters); + double spectralAngle = CalculateNormalizedSpectralAngle(decoylibrarySpectrum, scan.TheScan, psm,commonParameters); psm.SpectralAngle = spectralAngle; } } @@ -72,7 +72,7 @@ public static List GetDecoyLibrarySpectrumFromTargetByRevers /// /// Calculates the spectral angle, as described by Prosit ( https://www.nature.com/articles/s41592-019-0426-7 ). /// - public static double CalculateNormalizedSpectralAngle(List theoreticalLibraryIons, MsDataScan scan, CommonParameters commonParameters) + public static double CalculateNormalizedSpectralAngle(List theoreticalLibraryIons, MsDataScan scan, PeptideSpectralMatch psm, CommonParameters commonParameters) { double mzCutoff = 300; int fragmentNumberCutoff = 3; @@ -108,11 +108,14 @@ public static double CalculateNormalizedSpectralAngle(List t var test = new Product(libraryIon.NeutralTheoreticalProduct.ProductType, libraryIon.NeutralTheoreticalProduct.Terminus, libraryIon.NeutralTheoreticalProduct.NeutralMass, libraryIon.NeutralTheoreticalProduct.FragmentNumber, libraryIon.NeutralTheoreticalProduct.AminoAcidPosition, libraryIon.NeutralTheoreticalProduct.NeutralLoss); + if (!matchedIons.ContainsKey(libraryIon)) + { + matchedIons.Add(libraryIon, new MatchedFragmentIon(ref test, mz, experimentalIntensity, libraryIon.Charge)); + } - matchedIons.Add(libraryIon, new MatchedFragmentIon(ref test, mz, experimentalIntensity, libraryIon.Charge)); } } - + psm.LibraryMatchedFragments = matchedIons.Values.ToList(); // L2 norm double expNormalizer = Math.Sqrt(matchedIons.Sum(p => Math.Pow(p.Value.Intensity, 2))); double theorNormalizer = Math.Sqrt(theoreticalLibraryIons.Sum(p => Math.Pow(p.Intensity, 2))); @@ -132,6 +135,75 @@ public static double CalculateNormalizedSpectralAngle(List t return normalizedSpectralAngle; } + public static LibrarySpectrum ConvertingPsmsToConcensusSpectrum(List mutiplePsms) + { + Dictionary allIntensityBeforeNormalize = new Dictionary(); + Dictionary allIntensityAfterNormalize = new Dictionary(); + Dictionary allMz = new Dictionary(); + Dictionary allAverageMz = new Dictionary(); + var allIntensitySumForNormalize = new double[mutiplePsms.Count]; + //Dictionary[]> allIons = new Dictionary[]>(); + Dictionary numberOfEachIons = new Dictionary(); + List libraryIons = new List(); + for (int i = 0; i < mutiplePsms.Count; i++) + { + + var matchedIons = mutiplePsms[i].MatchedFragmentIons.Where(p => p.Charge == 1).ToList(); + for (int j = 0; j < matchedIons.Count; j++) + { + Product newKey = matchedIons[j].NeutralTheoreticalProduct; + if (!allIntensityBeforeNormalize.ContainsKey(newKey)) + { + var listOfIntensity = new double[mutiplePsms.Count]; + var listOfMz = new double[mutiplePsms.Count]; + listOfIntensity[i] = matchedIons[j].Intensity; + listOfMz[i] = matchedIons[j].Mz; + allIntensityBeforeNormalize.Add(newKey, listOfIntensity); + allMz.Add(newKey, listOfMz); + numberOfEachIons.Add(newKey, 1); + } + else + { + allIntensityBeforeNormalize[newKey][i] = matchedIons[j].Intensity; + allMz[newKey][i] = matchedIons[j].Mz; + int num = numberOfEachIons[newKey]; + numberOfEachIons[newKey] = num + 1; + } + } + } + + List commonIons = numberOfEachIons.Where(p => p.Value == mutiplePsms.Count).Select(q => q.Key).ToList(); + for (int k = 0; k < mutiplePsms.Count; k++) + { + double intensitySum = 0; + foreach (var x in commonIons) + { + intensitySum = intensitySum + allIntensityBeforeNormalize[x][k]; + } + allIntensitySumForNormalize[k] = intensitySum; + } + + foreach (var y in allIntensityBeforeNormalize) + { + for (int l = 0; l < y.Value.Length; l++) + { + double intensityBeforeNormalize = y.Value[l]; + y.Value[l] = intensityBeforeNormalize / allIntensitySumForNormalize[l]; + } + allIntensityAfterNormalize.Add(y.Key, y.Value.Where(p=>p>0).ToList().Average()); + } + var max = allIntensityAfterNormalize.Select(p => p.Value).Max(); + foreach (var eachMz in allMz) + { + Product temProduct = eachMz.Key; + libraryIons.Add(new MatchedFragmentIon(ref temProduct, eachMz.Value.Average(), allIntensityAfterNormalize[eachMz.Key]/ max, 1)); + } + + return new LibrarySpectrum(mutiplePsms[0].FullSequence, mutiplePsms.Select(p => p.ScanPrecursorMonoisotopicPeakMz).Average(), mutiplePsms[0].ScanPrecursorCharge, libraryIons, mutiplePsms.Select(p => p.ScanRetentionTime).Average(), false); + + + } + public static double CalculatePsmsNormalizedSpectralAngle(List standardSpectrum, List spectrum) { double mzCutoff = 300; @@ -262,6 +334,166 @@ public static double MatchedSpectraCompare(List standardSpec return score; } + public static List newAverageTwoSpectra(List spectraOne, List spectraTwo) + { + Dictionary averagedPeaksDictionary = new Dictionary(); + var averageTwoSpectraResult = new List(); + Dictionary spectraOneDictionary = new Dictionary(); + Dictionary spectraTwoDictionary = new Dictionary(); + + foreach (var ionOne in spectraOne) + { + var productWithChargeInOne = ionOne.NeutralTheoreticalProduct.ProductType.ToString() + ionOne.NeutralTheoreticalProduct.FragmentNumber + "^" + ionOne.Charge; + spectraOneDictionary.Add(productWithChargeInOne, ionOne); + } + foreach (var ionTwo in spectraTwo) + { + var productWithChargeInTwo = ionTwo.NeutralTheoreticalProduct.ProductType.ToString() + ionTwo.NeutralTheoreticalProduct.FragmentNumber + "^" + ionTwo.Charge; + spectraTwoDictionary.Add(productWithChargeInTwo, ionTwo); + } + var oneKeyList = spectraOneDictionary.Keys.ToList(); + var twoKeyList = spectraTwoDictionary.Keys.ToList(); + double intensitySum1 = 0; + double intensitySum2 = 0; + var commonKeyList = oneKeyList.Intersect(twoKeyList); + foreach (var common in commonKeyList) + { + intensitySum1 = intensitySum1 + spectraOneDictionary[common].Intensity; + intensitySum2 = intensitySum2 + spectraOneDictionary[common].Intensity; + } + List AllproductWithCharge = new List(); + AllproductWithCharge.AddRange(oneKeyList); + foreach (var x in twoKeyList) + { + if (!AllproductWithCharge.Contains(x)) + { + AllproductWithCharge.Add(x); + } + + } + + //Tolerance tolerance = new PpmTolerance(40); + foreach (var productWithCharge in AllproductWithCharge) + { + if (spectraOneDictionary.ContainsKey(productWithCharge) && spectraTwoDictionary.ContainsKey(productWithCharge)) + { + var newMz = (spectraOneDictionary[productWithCharge].Mz + spectraTwoDictionary[productWithCharge].Mz )/2; + var normInten1 = spectraOneDictionary[productWithCharge].Intensity / intensitySum1; + var normInten2 = spectraTwoDictionary[productWithCharge].Intensity / intensitySum2; + var newNorIntensity = (normInten1 + normInten2)/2 ; + Product product = spectraOneDictionary[productWithCharge].NeutralTheoreticalProduct; + var newIon = new MatchedFragmentIon(ref product, newMz, newNorIntensity, spectraOneDictionary[productWithCharge].Charge); + averageTwoSpectraResult.Add(newIon); + } + else if (spectraOneDictionary.ContainsKey(productWithCharge)) + { + Product productOne = spectraOneDictionary[productWithCharge].NeutralTheoreticalProduct; + var normInten1 = spectraOneDictionary[productWithCharge].Intensity / intensitySum1; + var oneIon = new MatchedFragmentIon(ref productOne, spectraOneDictionary[productWithCharge].Mz, normInten1, spectraOneDictionary[productWithCharge].Charge); + averageTwoSpectraResult.Add(oneIon); + } + else if (spectraTwoDictionary.ContainsKey(productWithCharge)) + { + Product productTwo = spectraTwoDictionary[productWithCharge].NeutralTheoreticalProduct; + //var normInten2 = spectraTwoDictionary[productWithCharge].Intensity / intensitySum2; + var normInten2 = spectraTwoDictionary[productWithCharge].Intensity / intensitySum2; + //var twoIon = new MatchedFragmentIon(ref productTwo, spectraTwoDictionary[productWithCharge].Mz, normInten2 / PsmsNum, spectraTwoDictionary[productWithCharge].Charge); + var twoIon = new MatchedFragmentIon(ref productTwo, spectraTwoDictionary[productWithCharge].Mz, normInten2, spectraTwoDictionary[productWithCharge].Charge); + averageTwoSpectraResult.Add(twoIon); + } + } + var testmz1 = averageTwoSpectraResult.Select(b => b.Mz).ToArray(); + var testin1 = averageTwoSpectraResult.Select(b => b.Intensity).ToArray(); + return averageTwoSpectraResult; + } + public static (List, Dictionary) newAverageTwoSpectra(List spectraOne, List spectraTwo, Dictionary ConsensusSpectraPeaksNumPair) + { + Dictionary averagedPeaksDictionary = new Dictionary(); + var averageTwoSpectraResult = new List(); + Dictionary spectraOneDictionary = new Dictionary(); + Dictionary spectraTwoDictionary = new Dictionary(); + //double intensitySum1 = spectraOne.Select(b => b.Intensity).Sum(); + //double intensitySum2 = spectraTwo.Select(b => b.Intensity).Sum(); + //double intensityMax1 = spectraOne.Select(b => b.Intensity).Max(); + //double intensityMax2 = spectraTwo.Select(b => b.Intensity).Max(); + foreach (var ionOne in spectraOne) + { + var productWithChargeInOne = ionOne.NeutralTheoreticalProduct.ProductType.ToString() + ionOne.NeutralTheoreticalProduct.FragmentNumber + "^" + ionOne.Charge; + spectraOneDictionary.Add(productWithChargeInOne, ionOne); + } + foreach (var ionTwo in spectraTwo) + { + var productWithChargeInTwo = ionTwo.NeutralTheoreticalProduct.ProductType.ToString() + ionTwo.NeutralTheoreticalProduct.FragmentNumber + "^" + ionTwo.Charge; + spectraTwoDictionary.Add(productWithChargeInTwo, ionTwo); + } + var oneKeyList = spectraOneDictionary.Keys.ToList(); + var twoKeyList = spectraTwoDictionary.Keys.ToList(); + double intensitySum1 = 0; + double intensitySum2 = 0; + var commonKeyList = oneKeyList.Intersect(twoKeyList); + foreach(var common in commonKeyList) + { + intensitySum1 = intensitySum1 + spectraOneDictionary[common].Intensity; + intensitySum2 = intensitySum2 + spectraOneDictionary[common].Intensity; + } + List AllproductWithCharge = new List(); + AllproductWithCharge.AddRange(oneKeyList); + foreach (var x in twoKeyList) + { + if (!AllproductWithCharge.Contains(x)) + { + AllproductWithCharge.Add(x); + } + + } + + //Tolerance tolerance = new PpmTolerance(40); + foreach (var productWithCharge in AllproductWithCharge) + { + if (spectraOneDictionary.ContainsKey(productWithCharge) && spectraTwoDictionary.ContainsKey(productWithCharge)) + { + if (ConsensusSpectraPeaksNumPair != null && ConsensusSpectraPeaksNumPair.ContainsKey(productWithCharge)) + { + ConsensusSpectraPeaksNumPair[productWithCharge] = ConsensusSpectraPeaksNumPair[productWithCharge] + 1; + } + else + { + ConsensusSpectraPeaksNumPair.Add(productWithCharge, 2); + } + int peaksNum = ConsensusSpectraPeaksNumPair[productWithCharge]; + var newMz = spectraOneDictionary[productWithCharge].Mz * (1 - (1.0 / peaksNum)) + spectraTwoDictionary[productWithCharge].Mz / peaksNum; + //var normInten1 = spectraOneDictionary[productWithCharge].Intensity / intensitySum1; + //var normInten2 = spectraTwoDictionary[productWithCharge].Intensity / intensitySum2; + var normInten1 = spectraOneDictionary[productWithCharge].Intensity / intensitySum1; + var normInten2 = spectraTwoDictionary[productWithCharge].Intensity / intensitySum2; + var newNorIntensity = normInten1 * (1 - (1.0 / peaksNum)) + normInten2 / peaksNum; + Product product = spectraOneDictionary[productWithCharge].NeutralTheoreticalProduct; + var newIon = new MatchedFragmentIon(ref product, newMz, newNorIntensity, spectraOneDictionary[productWithCharge].Charge); + averageTwoSpectraResult.Add(newIon); + } + else if (spectraOneDictionary.ContainsKey(productWithCharge)) + { + Product productOne = spectraOneDictionary[productWithCharge].NeutralTheoreticalProduct; + //var normInten1 = spectraOneDictionary[productWithCharge].Intensity / intensitySum1; + var normInten1 = spectraOneDictionary[productWithCharge].Intensity / intensitySum1; + //var oneIon = new MatchedFragmentIon(ref productOne, spectraOneDictionary[productWithCharge].Mz, normInten1 * (1 - (1.0 / PsmsNum)), spectraOneDictionary[productWithCharge].Charge); + var oneIon = new MatchedFragmentIon(ref productOne, spectraOneDictionary[productWithCharge].Mz, normInten1, spectraOneDictionary[productWithCharge].Charge); + averageTwoSpectraResult.Add(oneIon); + } + else if (spectraTwoDictionary.ContainsKey(productWithCharge)) + { + Product productTwo = spectraTwoDictionary[productWithCharge].NeutralTheoreticalProduct; + //var normInten2 = spectraTwoDictionary[productWithCharge].Intensity / intensitySum2; + var normInten2 = spectraTwoDictionary[productWithCharge].Intensity / intensitySum2; + //var twoIon = new MatchedFragmentIon(ref productTwo, spectraTwoDictionary[productWithCharge].Mz, normInten2 / PsmsNum, spectraTwoDictionary[productWithCharge].Charge); + var twoIon = new MatchedFragmentIon(ref productTwo, spectraTwoDictionary[productWithCharge].Mz, normInten2, spectraTwoDictionary[productWithCharge].Charge); + averageTwoSpectraResult.Add(twoIon); + } + } + var testmz1 = averageTwoSpectraResult.Select(b => b.Mz).ToArray(); + var testin1 = averageTwoSpectraResult.Select(b => b.Intensity).ToArray(); + return (averageTwoSpectraResult, ConsensusSpectraPeaksNumPair); + } public static (List, Dictionary) AverageTwoSpectra(List spectraOne, List spectraTwo, Dictionary ConsensusSpectraPeaksNumPair) { Dictionary averagedPeaksDictionary = new Dictionary(); diff --git a/MetaMorpheus.sln b/MetaMorpheus02.sln similarity index 100% rename from MetaMorpheus.sln rename to MetaMorpheus02.sln diff --git a/TaskLayer/MetaMorpheusTask.cs b/TaskLayer/MetaMorpheusTask.cs index 302f2e860..dd62bdd46 100644 --- a/TaskLayer/MetaMorpheusTask.cs +++ b/TaskLayer/MetaMorpheusTask.cs @@ -610,6 +610,43 @@ protected static void WriteSpectralLibrary(List spectrumLibrary } } + //protected static void WriteSpectralLibrary(Dictionary, List)> spectrumLibrary, string outputFolder) + //{ + + // string spectrumFilePath = outputFolder + "\\spectralLibrary.msp"; + // using (StreamWriter output = new StreamWriter(spectrumFilePath)) + // { + + + // foreach (var x in spectrumLibrary) + // { + // StringBuilder spectrum = new StringBuilder(); + // spectrum.Append("Name: "); + // spectrum.Append("\nMW: "); + // spectrum.Append("\nComment: "); + // spectrum.Append("Parent="); + // spectrum.Append(" RT="); + // spectrum.Append("\nNum peaks: " + spectrumLibrary.Count); + + // double maxIntensity = MatchedFragmentIons.Select(b => b.Intensity).Max(); + // double sumIntensity = MatchedFragmentIons.Select(b => b.Intensity).Sum(); + + // foreach (MatchedFragmentIon matchedIon in MatchedFragmentIons) + // { + // //double intensityFraction = matchedIon.Intensity / sumIntensity; + // double intensityFraction = matchedIon.Intensity / maxIntensity; + + // spectrum.Append("\n" + matchedIon.Mz + "\t" + intensityFraction + "\t" + "\"" + + // matchedIon.NeutralTheoreticalProduct.ProductType.ToString() + + // matchedIon.NeutralTheoreticalProduct.FragmentNumber.ToString() + "^" + + // matchedIon.Charge + "/" + 0 + "ppm" + "\""); + // } + // output.WriteLine(x.ToString()); + // } + + // } + //} + //forTestingLibraryGeneration protected static void WriteAllspectra(List spectrumLibrary, string outputFolder) { diff --git a/TaskLayer/SearchTask/PostSearchAnalysisParameters.cs b/TaskLayer/SearchTask/PostSearchAnalysisParameters.cs index f39b62641..1c537a0d5 100644 --- a/TaskLayer/SearchTask/PostSearchAnalysisParameters.cs +++ b/TaskLayer/SearchTask/PostSearchAnalysisParameters.cs @@ -25,5 +25,6 @@ public class PostSearchAnalysisParameters public MyFileManager MyFileManager { get; set; } public List DatabaseFilenameList { get; set; } public List CurrentRawFileList { get; set; } + public SpectralLibrary SpectralLibrary { get; set; } } } \ No newline at end of file diff --git a/TaskLayer/SearchTask/PostSearchAnalysisTask.cs b/TaskLayer/SearchTask/PostSearchAnalysisTask.cs index 413b17b85..2be951080 100644 --- a/TaskLayer/SearchTask/PostSearchAnalysisTask.cs +++ b/TaskLayer/SearchTask/PostSearchAnalysisTask.cs @@ -59,7 +59,7 @@ public MyTaskResults Run() .GroupBy(b => (b.FullFilePath, b.ScanNumber, b.PeptideMonisotopicMass)).Select(b => b.First()).ToList(); CalculatePsmFdr(); - } + } DoMassDifferenceLocalizationAnalysis(); ProteinAnalysis(); @@ -70,6 +70,11 @@ public MyTaskResults Run() HistogramAnalysis(); WritePsmResults(); SpectralLibraryGeneration(); + //SpectralLibraryGeneration(Parameters.SpectralLibrary); + if (Parameters.SpectralLibrary != null) + { + Parameters.SpectralLibrary.CloseConnections(); + } WriteProteinResults(); WriteQuantificationResults(); WritePrunedDatabase(); @@ -515,13 +520,15 @@ private void HistogramAnalysis() } } - //write spectral library by Yuling 11/14 + //write spectral library by Yuling 03/16 private void SpectralLibraryGeneration() { var FilteredPsmList = Parameters.AllPsms - .Where(p => p.FdrInfo.QValue <= 0.1 + .Where(p => p.FdrInfo.QValue <= 0.01 && p.FdrInfo.PEP < 0.5 && p.FdrInfo.QValueNotch <= CommonParameters.QValueOutputFilter).ToList(); - FilteredPsmList.RemoveAll(b => b.BestMatchingPeptides.First().Peptide.Protein.Accession.Contains("DECOY")); + //var FilteredPsmList = Parameters.AllPsms + // .Where(p => p.FdrInfo.QValueNotch <= CommonParameters.QValueOutputFilter).ToList(); + //FilteredPsmList.RemoveAll(b => b.BestMatchingPeptides.First().Peptide.Protein.Accession.Contains("DECOY")); FilteredPsmList.RemoveAll(b => b.IsDecoy); FilteredPsmList.RemoveAll(b => b.IsContaminant); @@ -529,7 +536,7 @@ private void SpectralLibraryGeneration() Dictionary> PsmsGroupByPeptideAndCharge = new Dictionary>(); foreach (var x in FilteredPsmList) { - List psmsWithsinglePeptide = Parameters.AllPsms.Where(b => b.FullSequence == x.FullSequence).OrderByDescending(p => p.Score).ToList(); + List psmsWithsinglePeptide = FilteredPsmList.Where(b => b.FullSequence == x.FullSequence).OrderByDescending(p => p.Score).ToList(); String peptideWithChargeState = x.FullSequence + "/" + x.ScanPrecursorCharge; List psmsWithsinglePeptideAndSameCharge = psmsWithsinglePeptide.Where(b => b.ScanPrecursorCharge == x.ScanPrecursorCharge).OrderByDescending(p => p.Score).ToList(); if (!PsmsGroupByPeptideAndCharge.ContainsKey(peptideWithChargeState)) @@ -541,61 +548,274 @@ private void SpectralLibraryGeneration() var spectrumLibrary = new List(); foreach (var x in PsmsGroupByPeptideAndCharge) { - List standspctra = x.Value[0].MatchedFragmentIons; - double standPrecursurMz = x.Value[0].ScanPrecursorMonoisotopicPeakMz; - double standRt = x.Value[0].ScanRetentionTime; - int a = 0; - var ConsensusSpectraPeaksNumPair = new Dictionary(); - int PsmsNum = 1; - while (a < x.Value.Count - 1) + List < PeptideSpectralMatch> psmsForLibrary = new List(); + if (x.Value.Count > 2) { - //Console.WriteLine(x.Key + " " + this.matchedSpectraCompare(standspctra, x.Value[a + 1].MatchedFragmentIons)); - var spectrumTocompare = x.Value[a + 1].MatchedFragmentIons; - //if (x.Value[0].FullSequence.Contains("HAEGTFTSDVSSYLEGQAAK")) + int a = 0; + int b = 0; + var ConsensusSpectraPeaksNumPair = new Dictionary(); + //int PsmsNum = 1; + List eachSaTotal = new List(); + while (b < 10 && b < x.Value.Count) + { + int c = 0; + List possibleStandard = x.Value[b].MatchedFragmentIons.Where(p => p.Charge == 1).ToList(); + List saTotal = new List(); + while (c < 10 && c < x.Value.Count) + { + var saOne = SpectralLibrarySearchFunction.CalculateNormalizedSpectralAngle(possibleStandard, x.Value[c].MsDataScan, x.Value[c], CommonParameters); + var sasaOne = SpectralLibrarySearchFunction.CalculatePsmsNormalizedSpectralAngle(possibleStandard.Where(p => p.Charge == 1).ToList(), x.Value[c].MatchedFragmentIons.Where(p => p.Charge == 1).ToList()); + var sasasaOne = SpectralLibrarySearchFunction.CalculatePsmsNormalizedSpectralAngle(x.Value[c].MatchedFragmentIons.Where(p => p.Charge == 1).ToList(), x.Value[c].MatchedFragmentIons.Where(p => p.Charge == 1).ToList()); + saTotal.Add(saOne); + c++; + } + eachSaTotal.Add(saTotal.Average()); + b++; + } + //if (eachSaTotal.Max() > 0.5) //{ + int standardIndex = eachSaTotal.IndexOf(eachSaTotal.Max()); + + List standspctra = x.Value[standardIndex].MatchedFragmentIons.Where(p => p.Charge == 1).ToList(); + psmsForLibrary.Add(x.Value[standardIndex]); + double standPrecursurMz = x.Value[standardIndex].ScanPrecursorMonoisotopicPeakMz; + double standRt = x.Value[standardIndex].ScanRetentionTime; + Dictionary, List)> allMatchedIons = new Dictionary, List)>(); + while (a < x.Value.Count - 1) + { + //Console.WriteLine(x.Key + " " + this.matchedSpectraCompare(standspctra, x.Value[a + 1].MatchedFragmentIons)); + var spectrumTocompare = x.Value[a].MatchedFragmentIons.Where(p => p.Charge == 1).ToList(); + //if (x.Value[0].FullSequence.Contains("TTGIVMDSGDGVTHTVPIYEGYALPHAILR")) + //{ + + //} + + //var compareScore = SpectralLibrarySearchFunction.MatchedSpectraCompare(standspctra, spectrumTocompare); + var testScore1 = SpectralLibrarySearchFunction.CalculatePsmsNormalizedSpectralAngle(x.Value[standardIndex].MatchedFragmentIons.Where(p => p.Charge == 1).ToList(), spectrumTocompare); + //var testscore2 = SpectralLibrarySearchFunction.CalculatePsmsNormalizedSpectralAngle(x.Value[standardIndex].MatchedFragmentIons.Where(p => p.Charge == 1).ToList(), spectrumTocompare.Where(p => p.Charge == 1).ToList()); + var testScore = SpectralLibrarySearchFunction.CalculateNormalizedSpectralAngle(standspctra, x.Value[a].MsDataScan, x.Value[a], CommonParameters); + //var sa = SpectralLibrarySearchFunction.CalculateNormalizedSpectralAngle(standspctra, x.Value[a + 1].MsDataScan, CommonParameters); + if (testScore > 0.67 && a != standardIndex) + { + psmsForLibrary.Add(x.Value[a]); + x.Value[a].ScoreForLibraryGeneration = testScore; + } + //if (testScore > 0.67 && a != standardIndex && a < 5) + //{ + // standspctra = SpectralLibrarySearchFunction.newAverageTwoSpectra(standspctra, x.Value[a].LibraryMatchedFragments); + // standPrecursurMz = (standPrecursurMz + x.Value[a].ScanPrecursorMonoisotopicPeakMz) / 2; + // standRt = (standRt + x.Value[a].ScanRetentionTime) / 2; + //} + a++; + } + //if(psmsForLibrary.Count>1||(psmsForLibrary.Count==1&& eachSaTotal.Max()>0.5)) + //{ + var standSpectrum = SpectralLibrarySearchFunction.ConvertingPsmsToConcensusSpectrum(psmsForLibrary); + //var standSpectrum = new LibrarySpectrum(x.Value[0].FullSequence, standPrecursurMz, x.Value[0].ScanPrecursorCharge, standspctra, standRt); + standSpectrum.BaseSequenceWithoutMods = x.Value[0].BaseSequence; + spectrumLibrary.Add(standSpectrum); //} + + } + //} - var compareScore = SpectralLibrarySearchFunction.MatchedSpectraCompare(standspctra, spectrumTocompare); - var testScore = SpectralLibrarySearchFunction.CalculatePsmsNormalizedSpectralAngle(standspctra, spectrumTocompare); - if (compareScore > 0.95) - { - PsmsNum++; - var pairResult = SpectralLibrarySearchFunction.AverageTwoSpectra(standspctra, spectrumTocompare, ConsensusSpectraPeaksNumPair); - standspctra = pairResult.Item1; - ConsensusSpectraPeaksNumPair = pairResult.Item2; - standPrecursurMz = standPrecursurMz * (1 - (1.0 / PsmsNum)) + x.Value[a + 1].ScanPrecursorMonoisotopicPeakMz / PsmsNum; - standRt = standRt * (1 - (1.0 / PsmsNum)) + x.Value[a + 1].ScanRetentionTime / PsmsNum; - } - a++; + } + + WriteSpectralLibrary(spectrumLibrary, Parameters.OutputFolder); + + //fortesting + //var allspectra = new List(); + //foreach (var x in PsmsGroupByPeptideAndCharge) + //{ + // if (x.Value.Count > 1) + // { + // foreach (var each in x.Value) + // { + // allspectra.Add(new LibrarySpectrum(each.FullSequence, each.ScanPrecursorMonoisotopicPeakMz, each.ScanPrecursorCharge, each.MatchedFragmentIons.Where(p => p.Charge == 1).ToList(), each.ScanRetentionTime)); + // } + // } + + //} + //WriteAllspectra(allspectra, Parameters.OutputFolder); + } + //write spectral library by Yuling 11/14 + private void SpectralLibraryGeneration(SpectralLibrary spectralLibrary) + { + var FilteredPsmList = Parameters.AllPsms + .Where(p => p.FdrInfo.QValue <= 0.01 + && p.FdrInfo.QValueNotch <= CommonParameters.QValueOutputFilter && p.SpectralAngle > 0.7 ).ToList(); + //var FilteredPsmList = Parameters.AllPsms + // .Where(p => p.FdrInfo.QValue <= 0.01 + // && p.FdrInfo.QValueNotch <= 0.0001 ).ToList(); + //FilteredPsmList.RemoveAll(b => b.BestMatchingPeptides.First().Peptide.Protein.Accession.Contains("DECOY")); + FilteredPsmList.RemoveAll(b => b.IsDecoy); + FilteredPsmList.RemoveAll(b => b.IsContaminant); + + + //write spectral library by Yuling 11/14 + Dictionary> PsmsGroupByPeptideAndCharge = new Dictionary>(); + foreach (var x in FilteredPsmList) + { + List psmsWithsinglePeptide = FilteredPsmList.Where(b => b.FullSequence == x.FullSequence).OrderByDescending(p => p.Score).ToList(); + String peptideWithChargeState = x.FullSequence + "/" + x.ScanPrecursorCharge; + List psmsWithsinglePeptideAndSameCharge = psmsWithsinglePeptide.Where(b => b.ScanPrecursorCharge == x.ScanPrecursorCharge).OrderBy(p => p.SpectralAngle).ToList(); + if (!PsmsGroupByPeptideAndCharge.ContainsKey(peptideWithChargeState)) + { + PsmsGroupByPeptideAndCharge.Add(peptideWithChargeState, psmsWithsinglePeptideAndSameCharge); } - foreach(var s in ConsensusSpectraPeaksNumPair) + } + + Dictionary spectrumLibrary = new Dictionary(); + + foreach (var x in PsmsGroupByPeptideAndCharge) + { + //string single_pdeep = @"g:\yuling\SLSpaper\testData\pdeepTTGIVMDSGDGVTHTVPIYEGYALPHAILR.msp"; + //var single_pdeepLibrary = new SpectralLibrary(new List { single_pdeep }); + //single_pdeepLibrary.TryGetSpectrum("TTGIVMDSGDGVTHTVPIYEGYALPHAILR", x.Value[0].ScanPrecursorCharge, out var pdeepSpectrum); + int aa = 0; + //var standSpectrum = new LibrarySpectrum(x.Value[0].FullSequence, x.Value[0].ScanPrecursorMonoisotopicPeakMz, x.Value[0].ScanPrecursorCharge, x.Value[0].LibraryMatchedFragments, x.Value[0].ScanRetentionTime); + var tempStand = x.Value[0].LibraryMatchedFragments; + double tempMz = x.Value[0].ScanPrecursorMonoisotopicPeakMz; + double temTime = x.Value[0].ScanRetentionTime; + while (aa < x.Value.Count - 1) { - if(s.Value < 0.6 * PsmsNum) - { - standspctra.RemoveAll(b => (b.NeutralTheoreticalProduct.ProductType.ToString() + b.NeutralTheoreticalProduct.FragmentNumber + "^" + b.Charge) == s.Key); - } + tempStand = SpectralLibrarySearchFunction.newAverageTwoSpectra(tempStand, x.Value[aa + 1].LibraryMatchedFragments); + //var test = SpectralLibrarySearchFunction.CalculatePsmsNormalizedSpectralAngle(pdeepSpectrum.MatchedFragmentIons, tempStand); + tempMz = (tempMz + x.Value[aa + 1].ScanPrecursorMonoisotopicPeakMz) / 2; + temTime = (temTime + x.Value[aa + 1].ScanRetentionTime) / 2; + aa++; } - var standSpectrum = new LibrarySpectrum(x.Value[0].FullSequence, standPrecursurMz, x.Value[0].ScanPrecursorCharge, standspctra, standRt); + var standSpectrum = new LibrarySpectrum(x.Value[0].FullSequence, tempMz, x.Value[0].ScanPrecursorCharge, tempStand, temTime); + //if (x.Value.Count == 1) + //{ + // var standSpectrum = new LibrarySpectrum(x.Value[0].FullSequence, x.Value[0].ScanPrecursorMonoisotopicPeakMz, x.Value[0].ScanPrecursorCharge, x.Value[0].MatchedFragmentIons, x.Value[0].ScanRetentionTime); + //} + //else if (x.Value.Count == 2) + //{ + // var standspctra = SpectralLibrarySearchFunction.newAverageTwoSpectra(x.Value[0].MatchedFragmentIons, x.Value[1].MatchedFragmentIons); + // var standSpectrum = new LibrarySpectrum(x.Value[0].FullSequence, x.Value[0].ScanPrecursorMonoisotopicPeakMz, x.Value[0].ScanPrecursorCharge, standspctra, x.Value[0].ScanRetentionTime); + //} + //else + //{ + // //List standspctra = x.Value[1].MatchedFragmentIons; + // //double standPrecursurMz = x.Value[0].ScanPrecursorMonoisotopicPeakMz; + // //double standRt = x.Value[0].ScanRetentionTime; + // int a = 0; + // int b = 0; + // var ConsensusSpectraPeaksNumPair = new Dictionary(); + // int PsmsNum = 1; + // List eachSaTotal = new List(); + // while (b < x.Value.Count - 1 && b < 5) + // { + // int c = 0; + // var possibleStandard = x.Value[b].MatchedFragmentIons; + // double saTotal = 0; + // while (c < x.Value.Count - 1 && c < 5) + // { + //var saOne = SpectralLibrarySearchFunction.CalculateNormalizedSpectralAngle(possibleStandard, x.Value[c].MsDataScan, CommonParameters); + // saTotal = saTotal + saOne; + // c++; + // } + // eachSaTotal.Add(saTotal); + // b++; + // } + // int standardIndex = eachSaTotal.IndexOf(eachSaTotal.Max()); + + // List standspctra = x.Value[standardIndex].MatchedFragmentIons; + // double standPrecursurMz = x.Value[standardIndex].ScanPrecursorMonoisotopicPeakMz; + // double standRt = x.Value[standardIndex].ScanRetentionTime; + // Dictionary, List)> allMatchedIons = new Dictionary, List)>(); + // while (a < x.Value.Count - 1) + // { + // //Console.WriteLine(x.Key + " " + this.matchedSpectraCompare(standspctra, x.Value[a + 1].MatchedFragmentIons)); + // var spectrumTocompare = x.Value[a].MatchedFragmentIons; + // //if (x.Value[0].FullSequence.Contains("HAEGTFTSDVSSYLEGQAAK")) + // //{ + + // //} + + // var compareScore = SpectralLibrarySearchFunction.MatchedSpectraCompare(standspctra, spectrumTocompare); + // var testScore = SpectralLibrarySearchFunction.CalculatePsmsNormalizedSpectralAngle(x.Value[standardIndex].MatchedFragmentIons, spectrumTocompare); + // var sa = SpectralLibrarySearchFunction.CalculateNormalizedSpectralAngle(standspctra, x.Value[a + 1].MsDataScan, CommonParameters); + // //if (testScore > 0.7 && a == standardIndex - 1) + // //{ + // // foreach (var eachIon in x.Value[a + 1].MatchedFragmentIons) + // // { + // // String newKey = eachIon.NeutralTheoreticalProduct.ProductType.ToString() + eachIon.NeutralTheoreticalProduct.FragmentNumber+"^" + eachIon.Charge; + // // if (!allMatchedIons.ContainsKey(newKey)) + // // { + // // var newIntensity = new List(); + // // var newMz = new List(); + // // newIntensity.Add(eachIon.Intensity); + // // newMz.Add(eachIon.Mz); + // // allMatchedIons.Add(newKey, (newIntensity, newMz)); + // // } + // // else + // // { + // // allMatchedIons[newKey].Item1.Add(eachIon.Intensity); + // // allMatchedIons[newKey].Item1.Add(eachIon.Mz); + // // } + // // } + // // PsmsNum++; + + // //} + + // //string single_pdeep = @"g:\yuling\SLSpaper\testData\single_pdeep.msp"; + // // var single_pdeepLibrary = new SpectralLibrary(new List { single_pdeep }); + // //single_pdeepLibrary.TryGetSpectrum("VTDEILHLVPNIDNFR", 3, out var pdeepSpectrum); + // if (testScore > 0.7 && a != standardIndex) + // { + // PsmsNum++; + // //var pairResult = SpectralLibrarySearchFunction.newAverageTwoSpectra(standspctra, spectrumTocompare, ConsensusSpectraPeaksNumPair); + // //standspctra = pairResult.Item1; + // //ConsensusSpectraPeaksNumPair = pairResult.Item2; + // standspctra = SpectralLibrarySearchFunction.newAverageTwoSpectra(standspctra, spectrumTocompare); + // //single_pdeepLibrary.TryGetSpectrum("VTDEILHLVPNIDNFR", 3, out var pdeepSpectrum); + // //var sss = SpectralLibrarySearchFunction.CalculatePsmsNormalizedSpectralAngle(pdeepSpectrum.MatchedFragmentIons, standspctra); + // //standPrecursurMz = standPrecursurMz * (1 - (1.0 / PsmsNum)) + x.Value[a + 1].ScanPrecursorMonoisotopicPeakMz / PsmsNum; + // //standRt = standRt * (1 - (1.0 / PsmsNum)) + x.Value[a + 1].ScanRetentionTime / PsmsNum; + // } + // a++; + // } + // //foreach(var s in ConsensusSpectraPeaksNumPair) + // //{ + // // if(s.Value < 0.6 * PsmsNum) + // // { + // // standspctra.RemoveAll(b => (b.NeutralTheoreticalProduct.ProductType.ToString() + b.NeutralTheoreticalProduct.FragmentNumber + "^" + b.Charge) == s.Key); + // // } + // //} + // var standSpectrum = new LibrarySpectrum(x.Value[0].FullSequence, standPrecursurMz, x.Value[0].ScanPrecursorCharge, standspctra, standRt); + //} standSpectrum.BaseSequenceWithoutMods = x.Value[0].BaseSequence; - spectrumLibrary.Add(standSpectrum); + spectrumLibrary.Add(x.Key, standSpectrum); } + var allSpectraInLibrary = spectralLibrary.GetAllLibrarySpectra(); + foreach (var librarySpectrum in allSpectraInLibrary) + { + String newKey = librarySpectrum.Sequence + "/" + librarySpectrum.ChargeState; + if (!spectrumLibrary.ContainsKey(newKey)) + { + spectrumLibrary.Add(newKey, librarySpectrum); + } - WriteSpectralLibrary(spectrumLibrary, Parameters.OutputFolder); + } + + WriteSpectralLibrary(spectrumLibrary.Values.ToList(), Parameters.OutputFolder); //fortesting //var allspectra = new List(); //foreach (var x in PsmsGroupByPeptideAndCharge) //{ - // if(x.Value.Count>1) + // if (x.Value.Count > 1) // { - // foreach(var each in x.Value) + // foreach (var each in x.Value) // { // allspectra.Add(new LibrarySpectrum(each.FullSequence, each.ScanPrecursorMonoisotopicPeakMz, each.ScanPrecursorCharge, each.MatchedFragmentIons, each.ScanRetentionTime)); // } // } - + //} //WriteAllspectra(allspectra, Parameters.OutputFolder); } diff --git a/TaskLayer/SearchTask/SearchTask.cs b/TaskLayer/SearchTask/SearchTask.cs index aa1773ea4..0d9153925 100644 --- a/TaskLayer/SearchTask/SearchTask.cs +++ b/TaskLayer/SearchTask/SearchTask.cs @@ -356,10 +356,10 @@ protected override MyTaskResults RunSpecific(string OutputFolder, List { taskId, "Individual Spectra Files" })); } - if (spectralLibrary != null) - { - spectralLibrary.CloseConnections(); - } + //if (spectralLibrary != null) + //{ + // spectralLibrary.CloseConnections(); + //} ReportProgress(new ProgressEventArgs(100, "Done with all searches!", new List { taskId, "Individual Spectra Files" })); @@ -388,7 +388,9 @@ protected override MyTaskResults RunSpecific(string OutputFolder, List