public static void TestClassicSearchEngineTopDown()
{
CommonParameters CommonParameters = new CommonParameters(
digestionParams: new DigestionParams(protease: "top-down"),
scoreCutoff: 1,
assumeOrphanPeaksAreZ1Fragments: false);
MetaMorpheusTask.DetermineAnalyteType(CommonParameters);
// test output file name (should be proteoform and not peptide)
Assert.That(GlobalVariables.AnalyteType == "Proteoform");
var variableModifications = new List <Modification>();
var fixedModifications = new List <Modification>();
var proteinList = new List <Protein>
{
new Protein("MPKVYSYQEVAEHNGPENFWIIIDDKVYDVSQFKDEHPGGDEIIMDLGGQDATESFVDIGHSDEALRLLKGLYIGDVDKTSERVSVEKVSTSENQSKGSGTLVVILAILMLGVAYYLLNE", "P40312")
};
var myMsDataFile = Mzml.LoadAllStaticData(Path.Combine(TestContext.CurrentContext.TestDirectory, @"TopDownTestData\slicedTDYeast.mzML"));
var searchMode = new SinglePpmAroundZeroSearchMode(5);
Tolerance DeconvolutionMassTolerance = new PpmTolerance(5);
var listOfSortedms2Scans = MetaMorpheusTask.GetMs2Scans(myMsDataFile, null, new CommonParameters()).OrderBy(b => b.PrecursorMass).ToArray();
PeptideSpectralMatch[] allPsmsArray = new PeptideSpectralMatch[listOfSortedms2Scans.Length];
new ClassicSearchEngine(allPsmsArray, listOfSortedms2Scans, variableModifications, fixedModifications, null, null, null,
proteinList, searchMode, CommonParameters, null, null, new List <string>()).Run();
var psm = allPsmsArray.Where(p => p != null).FirstOrDefault();
Assert.That(psm.MatchedFragmentIons.Count == 47);
}
public static void TestLookingForAcceptableIsotopicEnvelopes()
{
CommonParameters CommonParameters = new CommonParameters();
MetaMorpheusTask.DetermineAnalyteType(CommonParameters);
var variableModifications = new List <Modification>();
var fixedModifications = new List <Modification>();
var proteinList = new List <Protein>
{
new Protein("AAAHSSLK", ""), new Protein("RQPAQPR", ""), new Protein("EKAEAEAEK", "")
};
var myMsDataFile = Mzml.LoadAllStaticData(Path.Combine(TestContext.CurrentContext.TestDirectory, @"TestData\SmallCalibratible_Yeast.mzML"));
var searchMode = new SinglePpmAroundZeroSearchMode(5);
Tolerance DeconvolutionMassTolerance = new PpmTolerance(5);
var listOfSortedms2Scans = MetaMorpheusTask.GetMs2Scans(myMsDataFile, null, new CommonParameters()).OrderBy(b => b.PrecursorMass).ToArray();
var ms2ScanTest = listOfSortedms2Scans[0];
//test when all the masses are not in the given range
//test1 when all the masses are too small
var test1 = ms2ScanTest.GetClosestExperimentalIsotopicEnvelopeList(50, 95);
Assert.AreEqual(test1, null);
//test2 when all the masses are too big
var test2 = ms2ScanTest.GetClosestExperimentalIsotopicEnvelopeList(582, 682);
Assert.AreEqual(test2, null);
//test3 when the mass which is bigger than given min mass is bigger than the mass which is smaller than the given max mass
//for example: the mass array is [1,2,3,4,5], the given min mass is 2.2, the given max mass is 2.8
var test3 = ms2ScanTest.GetClosestExperimentalIsotopicEnvelopeList(110, 111);
Assert.AreEqual(test3, null);
//test normal conditions:look for IsotopicEnvelopes which are in the range of acceptable mass
var test4 = ms2ScanTest.GetClosestExperimentalIsotopicEnvelopeList(120, 130);
IsotopicEnvelope[] expected4 = ms2ScanTest.ExperimentalFragments.Skip(15).Take(9).ToArray();
Assert.That(ms2ScanTest.ExperimentalFragments[15].MonoisotopicMass > 120 && ms2ScanTest.ExperimentalFragments[14].MonoisotopicMass < 120);
Assert.That(ms2ScanTest.ExperimentalFragments[23].MonoisotopicMass < 130 && ms2ScanTest.ExperimentalFragments[24].MonoisotopicMass > 130);
Assert.AreEqual(test4, expected4);
var test5 = ms2ScanTest.GetClosestExperimentalIsotopicEnvelopeList(400, 500);
IsotopicEnvelope[] expected5 = ms2ScanTest.ExperimentalFragments.Skip(150).Take(7).ToArray();
Assert.That(ms2ScanTest.ExperimentalFragments[150].MonoisotopicMass > 400 && ms2ScanTest.ExperimentalFragments[149].MonoisotopicMass < 400);
Assert.That(ms2ScanTest.ExperimentalFragments[156].MonoisotopicMass < 500 && ms2ScanTest.ExperimentalFragments[157].MonoisotopicMass > 500);
Assert.AreEqual(test5, expected5);
}
public static void TestReverseDecoyGenerationDuringSearch()
{
CommonParameters CommonParameters = new CommonParameters();
MetaMorpheusTask.DetermineAnalyteType(CommonParameters);
var variableModifications = new List <Modification>();
var fixedModifications = new List <Modification>();
var proteinList = new List <Protein>
{
new Protein("KKAEDGINK", ""), new Protein("AVNSISLK", ""), new Protein("EKAEAEAEK", ""), new Protein("DITANLR", ""), new Protein("QNAIGTAK", ""),
new Protein("FHKSQLNK", ""), new Protein("KQVAQWNK", ""), new Protein("NTRIEELK", ""), new Protein("RQPAQPR", ""),
};
var myMsDataFile = Mzml.LoadAllStaticData(Path.Combine(TestContext.CurrentContext.TestDirectory, @"TestData\SmallCalibratible_Yeast.mzML"));
var searchMode = new SinglePpmAroundZeroSearchMode(5);
Tolerance DeconvolutionMassTolerance = new PpmTolerance(5);
var listOfSortedms2Scans = MetaMorpheusTask.GetMs2Scans(myMsDataFile, null, new CommonParameters()).OrderBy(b => b.PrecursorMass).ToArray();
var path = Path.Combine(TestContext.CurrentContext.TestDirectory, @"TestData\myPrositLib.msp");
var testLibrary = new SpectralLibrary(new List <string> {
path
});
//test when doing spectral library search without generating library
PeptideSpectralMatch[] allPsmsArray1 = new PeptideSpectralMatch[listOfSortedms2Scans.Length];
new ClassicSearchEngine(allPsmsArray1, listOfSortedms2Scans, variableModifications, fixedModifications, null, null, null,
proteinList, searchMode, CommonParameters, null, testLibrary, new List <string>(), false).Run();
var psm1 = allPsmsArray1.Where(p => p != null).ToList();
Assert.That(psm1[0].IsDecoy == false && psm1[0].FullSequence == "DITANLR");
Assert.That(psm1[1].IsDecoy == true && psm1[1].FullSequence == "LSISNVAK");
Assert.That(psm1[2].IsDecoy == true && psm1[2].FullSequence == "LSISNVAK");
Assert.That(psm1[3].IsDecoy == false && psm1[3].FullSequence == "RQPAQPR");
Assert.That(psm1[4].IsDecoy == false && psm1[4].FullSequence == "KKAEDGINK");
Assert.That(psm1[5].IsDecoy == false && psm1[5].FullSequence == "EKAEAEAEK");
Assert.That(psm1[6].IsDecoy == false && psm1[6].FullSequence == "EKAEAEAEK");
proteinList.Add(new Protein("LSISNVAK", "", isDecoy: true));
//test when doing spectral library search with generating library; non spectral search won't generate decoy by "decoy on the fly" , so proteinlist used by non spectral library search would contain decoys
PeptideSpectralMatch[] allPsmsArray2 = new PeptideSpectralMatch[listOfSortedms2Scans.Length];
new ClassicSearchEngine(allPsmsArray2, listOfSortedms2Scans, variableModifications, fixedModifications, null, null, null,
proteinList, searchMode, CommonParameters, null, testLibrary, new List <string>(), true).Run();
var psm2 = allPsmsArray2.Where(p => p != null).ToList();
Assert.That(psm2[0].IsDecoy == false && psm2[0].FullSequence == "DITANLR");
Assert.That(psm2[1].IsDecoy == true && psm2[1].FullSequence == "LSISNVAK");
Assert.That(psm2[2].IsDecoy == true && psm2[2].FullSequence == "LSISNVAK");
Assert.That(psm2[3].IsDecoy == false && psm2[3].FullSequence == "RQPAQPR");
Assert.That(psm2[4].IsDecoy == false && psm2[4].FullSequence == "KKAEDGINK");
Assert.That(psm2[5].IsDecoy == false && psm2[5].FullSequence == "EKAEAEAEK");
Assert.That(psm2[6].IsDecoy == false && psm2[6].FullSequence == "EKAEAEAEK");
//test when doing non spectral library search without generating library
PeptideSpectralMatch[] allPsmsArray3 = new PeptideSpectralMatch[listOfSortedms2Scans.Length];
new ClassicSearchEngine(allPsmsArray3, listOfSortedms2Scans, variableModifications, fixedModifications, null, null, null,
proteinList, searchMode, CommonParameters, null, null, new List <string>(), false).Run();
var psm3 = allPsmsArray3.Where(p => p != null).ToList();
Assert.That(psm3[0].IsDecoy == false && psm3[0].FullSequence == "DITANLR");
Assert.That(psm3[1].IsDecoy == true && psm3[1].FullSequence == "LSISNVAK");
Assert.That(psm3[2].IsDecoy == true && psm3[2].FullSequence == "LSISNVAK");
Assert.That(psm3[3].IsDecoy == false && psm3[3].FullSequence == "RQPAQPR");
Assert.That(psm3[4].IsDecoy == false && psm3[4].FullSequence == "KKAEDGINK");
Assert.That(psm3[5].IsDecoy == false && psm3[5].FullSequence == "EKAEAEAEK");
Assert.That(psm3[6].IsDecoy == false && psm3[6].FullSequence == "EKAEAEAEK");
//test when doing non spectral library search with generating library
PeptideSpectralMatch[] allPsmsArray4 = new PeptideSpectralMatch[listOfSortedms2Scans.Length];
new ClassicSearchEngine(allPsmsArray4, listOfSortedms2Scans, variableModifications, fixedModifications, null, null, null,
proteinList, searchMode, CommonParameters, null, null, new List <string>(), true).Run();
var psm4 = allPsmsArray4.Where(p => p != null).ToList();
Assert.That(psm4[0].IsDecoy == false && psm4[0].FullSequence == "DITANLR");
Assert.That(psm4[1].IsDecoy == true && psm4[1].FullSequence == "LSISNVAK");
Assert.That(psm4[2].IsDecoy == true && psm4[2].FullSequence == "LSISNVAK");
Assert.That(psm4[3].IsDecoy == false && psm4[3].FullSequence == "RQPAQPR");
Assert.That(psm4[4].IsDecoy == false && psm4[4].FullSequence == "KKAEDGINK");
Assert.That(psm4[5].IsDecoy == false && psm4[5].FullSequence == "EKAEAEAEK");
Assert.That(psm4[6].IsDecoy == false && psm4[6].FullSequence == "EKAEAEAEK");
//compare psm's target/decoy results in 4 conditions. they should be same as new decoy methods shouldn't change the t/d results
for (int i = 0; i < psm1.Count; i++)
{
Assert.That(psm1[i].FullSequence == psm2[i].FullSequence && psm3[i].FullSequence == psm3[i].FullSequence && psm2[i].FullSequence == psm3[i].FullSequence);
Assert.That(psm1[i].IsDecoy == psm2[i].IsDecoy && psm3[i].IsDecoy == psm3[i].IsDecoy && psm2[i].IsDecoy == psm3[i].IsDecoy);
}
//compare MetaMorpheus scores in 4 conditions; for some psms, they should have a little higher score when "generating library" as they switch to all charges ions matching function
for (int j = 0; j < psm1.Count; j++)
{
if (psm1[j].FullSequence == psm2[j].FullSequence && psm1[j].MatchedFragmentIons.Count != psm2[j].MatchedFragmentIons.Count)
{
Assert.That(psm1[j].Score < psm2[j].Score);
}
}
}
public static void TestMatchIonsOfAllChargesBottomUp()
{
CommonParameters CommonParameters = new CommonParameters();
MetaMorpheusTask.DetermineAnalyteType(CommonParameters);
var variableModifications = new List <Modification>();
var fixedModifications = new List <Modification>();
var proteinList = new List <Protein>
{
new Protein("AAAHSSLK", ""), new Protein("RQPAQPR", ""), new Protein("EKAEAEAEK", "")
};
var myMsDataFile = Mzml.LoadAllStaticData(Path.Combine(TestContext.CurrentContext.TestDirectory, @"TestData\SmallCalibratible_Yeast.mzML"));
var searchMode = new SinglePpmAroundZeroSearchMode(5);
Tolerance DeconvolutionMassTolerance = new PpmTolerance(5);
var listOfSortedms2Scans = MetaMorpheusTask.GetMs2Scans(myMsDataFile, null, new CommonParameters()).OrderBy(b => b.PrecursorMass).ToArray();
//search by new method of looking for all charges
PeptideSpectralMatch[] allPsmsArray = new PeptideSpectralMatch[listOfSortedms2Scans.Length];
new ClassicSearchEngine(allPsmsArray, listOfSortedms2Scans, variableModifications, fixedModifications, null, null, null,
proteinList, searchMode, CommonParameters, null, null, new List <string>(), true).Run();
var psm = allPsmsArray.Where(p => p != null).ToList();
Assert.That(psm[1].MatchedFragmentIons.Count == 14);
//there are ions with same product type and same fragment number but different charges
Assert.That(psm[1].MatchedFragmentIons[8].NeutralTheoreticalProduct.ProductType == psm[1].MatchedFragmentIons[9].NeutralTheoreticalProduct.ProductType &&
psm[1].MatchedFragmentIons[8].NeutralTheoreticalProduct.FragmentNumber == psm[1].MatchedFragmentIons[9].NeutralTheoreticalProduct.FragmentNumber &&
psm[1].MatchedFragmentIons[8].Charge != psm[1].MatchedFragmentIons[9].Charge);
Assert.That(psm[2].MatchedFragmentIons.Count == 14);
Assert.That(psm[4].MatchedFragmentIons.Count == 16);
//search by old method of looking for only one charge
PeptideSpectralMatch[] allPsmsArray_oneCharge = new PeptideSpectralMatch[listOfSortedms2Scans.Length];
new ClassicSearchEngine(allPsmsArray_oneCharge, listOfSortedms2Scans, variableModifications, fixedModifications, null, null, null,
proteinList, searchMode, CommonParameters, null, null, new List <string>(), false).Run();
var psm_oneCharge = allPsmsArray_oneCharge.Where(p => p != null).ToList();
//compare 2 scores , they should have same integer part but new search has a little higher score than old search
Assert.That(psm[1].Score > psm_oneCharge[1].Score);
Assert.AreEqual(Math.Truncate(psm[1].Score), 12);
Assert.AreEqual(Math.Truncate(psm_oneCharge[1].Score), 12);
//compare 2 results and evaluate the different matched ions
var peptideTheorProducts = new List <Product>();
Assert.That(psm_oneCharge[1].MatchedFragmentIons.Count == 12);
var differences = psm[1].MatchedFragmentIons.Except(psm_oneCharge[1].MatchedFragmentIons);
psm[1].BestMatchingPeptides.First().Peptide.Fragment(CommonParameters.DissociationType, CommonParameters.DigestionParams.FragmentationTerminus, peptideTheorProducts);
foreach (var ion in differences)
{
foreach (var product in peptideTheorProducts)
{
if (product.Annotation.ToString().Equals(ion.NeutralTheoreticalProduct.Annotation.ToString()))
{
//to see if the different matched ions are qualified
Assert.That(CommonParameters.ProductMassTolerance.Within(ion.Mz.ToMass(ion.Charge), product.NeutralMass));
}
}
}
//test specific condition: unknown fragment mass; this only happens rarely for sequences with unknown amino acids
var myMsDataFile1 = new TestDataFile();
var variableModifications1 = new List <Modification>();
var fixedModifications1 = new List <Modification>();
var proteinList1 = new List <Protein> {
new Protein("QXQ", null)
};
var productMassTolerance = new AbsoluteTolerance(0.01);
var searchModes = new OpenSearchMode();
Tolerance DeconvolutionMassTolerance1 = new PpmTolerance(5);
var listOfSortedms2Scans1 = MetaMorpheusTask.GetMs2Scans(myMsDataFile, null, new CommonParameters()).OrderBy(b => b.PrecursorMass).ToArray();
List <DigestionMotif> motifs = new List <DigestionMotif> {
new DigestionMotif("K", null, 1, null)
};
Protease protease = new Protease("Custom Protease3", CleavageSpecificity.Full, null, null, motifs);
ProteaseDictionary.Dictionary.Add(protease.Name, protease);
CommonParameters CommonParameters1 = new CommonParameters(
digestionParams: new DigestionParams(protease: protease.Name, maxMissedCleavages: 0, minPeptideLength: 1),
scoreCutoff: 1,
addCompIons: false);
var fsp = new List <(string fileName, CommonParameters fileSpecificParameters)>();
fsp.Add(("", CommonParameters));
PeptideSpectralMatch[] allPsmsArray1 = new PeptideSpectralMatch[listOfSortedms2Scans.Length];
bool writeSpectralLibrary = true;
new ClassicSearchEngine(allPsmsArray1, listOfSortedms2Scans1, variableModifications1, fixedModifications1, null, null, null,
proteinList1, searchModes, CommonParameters1, fsp, null, new List <string>(), writeSpectralLibrary).Run();
var psm1 = allPsmsArray1.Where(p => p != null).ToList();
Assert.AreEqual(psm1.Count, 222);
}
public static void TestMatchIonsOfAllChargesTopDown()
{
CommonParameters CommonParameters = new CommonParameters(
digestionParams: new DigestionParams(protease: "top-down"),
scoreCutoff: 1,
assumeOrphanPeaksAreZ1Fragments: false);
MetaMorpheusTask.DetermineAnalyteType(CommonParameters);
// test output file name (should be proteoform and not peptide)
Assert.That(GlobalVariables.AnalyteType == "Proteoform");
var variableModifications = new List <Modification>();
var fixedModifications = new List <Modification>();
var proteinList = new List <Protein>
{
new Protein("MPKVYSYQEVAEHNGPENFWIIIDDKVYDVSQFKDEHPGGDEIIMDLGGQDATESFVDIGHSDEALRLLKGLYIGDVDKTSERVSVEKVSTSENQSKGSGTLVVILAILMLGVAYYLLNE", "P40312")
};
var myMsDataFile = Mzml.LoadAllStaticData(Path.Combine(TestContext.CurrentContext.TestDirectory, @"TopDownTestData\slicedTDYeast.mzML"));
var searchMode = new SinglePpmAroundZeroSearchMode(5);
Tolerance DeconvolutionMassTolerance = new PpmTolerance(5);
var listOfSortedms2Scans = MetaMorpheusTask.GetMs2Scans(myMsDataFile, null, new CommonParameters()).OrderBy(b => b.PrecursorMass).ToArray();
//search by new method of looking for all charges
PeptideSpectralMatch[] allPsmsArray = new PeptideSpectralMatch[listOfSortedms2Scans.Length];
new ClassicSearchEngine(allPsmsArray, listOfSortedms2Scans, variableModifications, fixedModifications, null, null, null,
proteinList, searchMode, CommonParameters, null, null, new List <string>(), true).Run();
var psm = allPsmsArray.Where(p => p != null).FirstOrDefault();
Assert.That(psm.MatchedFragmentIons.Count == 62);
//search by old method of looking for only one charge
PeptideSpectralMatch[] allPsmsArray_oneCharge = new PeptideSpectralMatch[listOfSortedms2Scans.Length];
new ClassicSearchEngine(allPsmsArray_oneCharge, listOfSortedms2Scans, variableModifications, fixedModifications, null, null, null,
proteinList, searchMode, CommonParameters, null, null, new List <string>(), false).Run();
var psm_oneCharge = allPsmsArray_oneCharge.Where(p => p != null).FirstOrDefault();
Assert.That(psm_oneCharge.MatchedFragmentIons.Count == 47);
//compare 2 scores , they should have same integer but new search has a little higher score than old search
Assert.That(psm.Score > psm_oneCharge.Score);
Assert.AreEqual(Math.Truncate(psm.Score), 47);
Assert.AreEqual(Math.Truncate(psm_oneCharge.Score), 47);
//compare 2 results and evaluate the different matched ions
var peptideTheorProducts = new List <Product>();
var differences = psm.MatchedFragmentIons.Except(psm_oneCharge.MatchedFragmentIons);
psm.BestMatchingPeptides.First().Peptide.Fragment(CommonParameters.DissociationType, CommonParameters.DigestionParams.FragmentationTerminus, peptideTheorProducts);
foreach (var ion in differences)
{
foreach (var product in peptideTheorProducts)
{
if (product.Annotation.ToString().Equals(ion.NeutralTheoreticalProduct.Annotation.ToString()))
{
//to see if the different matched ions are qualified
Assert.That(CommonParameters.ProductMassTolerance.Within(ion.Mz.ToMass(ion.Charge), product.NeutralMass));
}
}
}
}