public static void TestCoIsolation()
{
List <DigestionMotif> motifs = new List <DigestionMotif> {
new DigestionMotif("K", null, 1, null)
};
Protease protease = new Protease("CustProtease", CleavageSpecificity.Full, null, null, motifs);
ProteaseDictionary.Dictionary.Add(protease.Name, protease);
CommonParameters CommonParameters = new CommonParameters(scoreCutoff: 1, deconvolutionIntensityRatio: 50, digestionParams: new DigestionParams(protease.Name, minPeptideLength: 1));
var variableModifications = new List <Modification>();
var fixedModifications = new List <Modification>();
var proteinList = new List <Protein> {
new Protein("MNNNKNDNK", null)
};
var searchModes = new SinglePpmAroundZeroSearchMode(5);
Proteomics.AminoAcidPolymer.Peptide pep1 = new Proteomics.AminoAcidPolymer.Peptide("NNNK");
Proteomics.AminoAcidPolymer.Peptide pep2 = new Proteomics.AminoAcidPolymer.Peptide("NDNK");
var dist1 = IsotopicDistribution.GetDistribution(pep1.GetChemicalFormula(), 0.1, 0.01);
var dist2 = IsotopicDistribution.GetDistribution(pep2.GetChemicalFormula(), 0.1, 0.01);
MsDataScan[] Scans = new MsDataScan[2];
double[] ms1intensities = new double[] { 0.8, 0.8, 0.2, 0.02, 0.2, 0.02 };
double[] ms1mzs = dist1.Masses.Concat(dist2.Masses).OrderBy(b => b).Select(b => b.ToMz(1)).ToArray();
double selectedIonMz = ms1mzs[1];
MzSpectrum MS1 = new MzSpectrum(ms1mzs, ms1intensities, false);
Scans[0] = new MsDataScan(MS1, 1, 1, false, Polarity.Positive, 1.0, new MzRange(300, 2000), "first spectrum", MZAnalyzerType.Unknown, MS1.SumOfAllY, null, null, "scan=1");
double[] ms2intensities = new double[] { 1, 1, 1, 1, 1 };
double[] ms2mzs = new double[] { 146.106.ToMz(1), 228.086.ToMz(1), 229.07.ToMz(1), 260.148.ToMz(1), 342.129.ToMz(1) };
MzSpectrum MS2 = new MzSpectrum(ms2mzs, ms2intensities, false);
double isolationMZ = selectedIonMz;
Scans[1] = new MsDataScan(MS2, 2, 2, false, Polarity.Positive, 2.0, new MzRange(100, 1500), "second spectrum", MZAnalyzerType.Unknown, MS2.SumOfAllY, null, null, "scan=2", selectedIonMz, null, null, isolationMZ, 2.5, DissociationType.HCD, 1, null);
var myMsDataFile = new MsDataFile(Scans, null);
var listOfSortedms2Scans = MetaMorpheusTask.GetMs2Scans(myMsDataFile, null, new CommonParameters(deconvolutionIntensityRatio: 50)).OrderBy(b => b.PrecursorMass).ToArray();
PeptideSpectralMatch[] allPsmsArray = new PeptideSpectralMatch[listOfSortedms2Scans.Length];;
new ClassicSearchEngine(allPsmsArray, listOfSortedms2Scans, variableModifications, fixedModifications, null, null, null, proteinList, searchModes, CommonParameters, new List <string>()).Run();
// Two matches for this single scan! Corresponding to two co-isolated masses
Assert.AreEqual(2, allPsmsArray.Length);
Assert.IsTrue(allPsmsArray[0].Score > 1);
Assert.AreEqual(2, allPsmsArray[0].ScanNumber);
Assert.AreEqual("NNNK", allPsmsArray[0].BaseSequence);
Assert.AreEqual("NDNK", allPsmsArray[1].BaseSequence);
}
public static void TestPeakSplittingRight()
{
string fileToWrite = "myMzml.mzML";
string peptide = "PEPTIDE";
double intensity = 1e6;
Loaders.LoadElements(Path.Combine(TestContext.CurrentContext.TestDirectory, @"elements.dat"));
// generate mzml file
// 1 MS1 scan per peptide
MsDataScan[] scans = new MsDataScan[10];
double[] intensityMultipliers = { 1, 3, 5, 10, 5, 3, 1, 1, 3, 1 };
for (int s = 0; s < scans.Length; s++)
{
ChemicalFormula cf = new Proteomics.AminoAcidPolymer.Peptide(peptide).GetChemicalFormula();
IsotopicDistribution dist = IsotopicDistribution.GetDistribution(cf, 0.125, 1e-8);
double[] mz = dist.Masses.Select(v => v.ToMz(1)).ToArray();
double[] intensities = dist.Intensities.Select(v => v * intensity * intensityMultipliers[s]).ToArray();
// add the scan
scans[s] = new MsDataScan(massSpectrum: new MzSpectrum(mz, intensities, false), oneBasedScanNumber: s + 1, msnOrder: 1, isCentroid: true,
polarity: Polarity.Positive, retentionTime: 1.0 + s / 10.0, scanWindowRange: new MzRange(400, 1600), scanFilter: "f",
mzAnalyzer: MZAnalyzerType.Orbitrap, totalIonCurrent: intensities.Sum(), injectionTime: 1.0, noiseData: null, nativeId: "scan=" + (s + 1));
}
// write the .mzML
IO.MzML.MzmlMethods.CreateAndWriteMyMzmlWithCalibratedSpectra(new FakeMsDataFile(scans),
Path.Combine(TestContext.CurrentContext.TestDirectory, fileToWrite), false);
// set up spectra file info
SpectraFileInfo file1 = new SpectraFileInfo(Path.Combine(TestContext.CurrentContext.TestDirectory, fileToWrite), "", 0, 0, 0);
// create some PSMs
var pg = new ProteinGroup("MyProtein", "gene", "org");
Identification id1 = new Identification(file1, peptide, peptide,
new Proteomics.AminoAcidPolymer.Peptide(peptide).MonoisotopicMass, 1.3 + 0.001, 1, new List <ProteinGroup> {
pg
});
// create the FlashLFQ engine
FlashLFQEngine engine = new FlashLFQEngine(new List <Identification> {
id1
});
// run the engine
var results = engine.Run();
ChromatographicPeak peak = results.Peaks.First().Value.First();
Assert.That(peak.Apex.RetentionTime == 1.3);
Assert.That(peak.SplitRT == 1.6);
Assert.That(!peak.IsotopicEnvelopes.Any(p => p.RetentionTime > 1.6));
Assert.That(peak.IsotopicEnvelopes.Count == 6);
}
public static double CalculateProteoformMass(string sequence, List <Ptm> ptm_combination)
{
if (Sweet.lollipop.theoretical_database.aaIsotopeMassList == null)
{
Sweet.lollipop.theoretical_database.populate_aa_mass_dictionary();
}
if (!Sweet.lollipop.use_average_mass)
{
double proteoformMass = 18.010565; // start with water
char[] aminoAcids = sequence.ToCharArray();
List <double> aaMasses = new List <double>();
for (int i = 0; i < sequence.Length; i++)
{
if (Sweet.lollipop.theoretical_database.aaIsotopeMassList.ContainsKey(aminoAcids[i]))
{
aaMasses.Add(Sweet.lollipop.theoretical_database.aaIsotopeMassList[aminoAcids[i]]);
}
}
return(proteoformMass + aaMasses.Sum() +
ptm_combination.Sum(p => Math.Round((double)p.modification.MonoisotopicMass, 5)));
}
//if most abundant mass, calculate iso distrubution, set modified/unmodified masses to most abundant.
var formula = new Proteomics.AminoAcidPolymer.Peptide(sequence).GetChemicalFormula();
// append mod formulas
foreach (var mod in ptm_combination)
{
var modCf = mod.modification.ChemicalFormula;
if (modCf != null)
{
formula.Add(modCf);
}
}
//Calculate isotopic distribution of the full peptide
//var dist = IsotopicDistribution.GetDistribution(formula, 0.1, 1e-12);
//double[] masses = dist.Masses.ToArray();
//double[] intensities = dist.Intensities.ToArray();
//double max = intensities.Max();
//int modeMassIndex = Array.IndexOf(intensities, max);
//return masses[modeMassIndex];
return(formula.AverageMass);
}
public ChemicalFormula GetChemicalFormula()
{
var formula = new Proteomics.AminoAcidPolymer.Peptide(sequence).GetChemicalFormula();
// append mod formulas
foreach (var mod in ptm_list)
{
var modCf = mod.modification.ChemicalFormula;
if (modCf != null)
{
formula.Add(modCf);
}
else
{
return(null);
}
}
return(formula);
}
public static void TestDeconvolution()
{
UsefulProteomicsDatabases.Loaders.LoadElements();
// >sp|P49703|ARL4D_HUMAN ADP-ribosylation factor-like protein 4D OS=H**o sapiens OX=9606 GN=ARL4D PE=1 SV=2
string sequence = "MGNHLTEMAPTASSFLPHFQALHVVVIGLDSAGKTSLLYRLKFKEFVQSVPTKGFNTEKIRVPLGGSRGITFQ" +
"VWDVGGQEKLRPLWRSYTRRTDGLVFVVDAAEAERLEEAKVELHRISRASDNQGVPVLVLANKQDQPGALSAA" +
"EVEKRLAVRELAAATLTHVQGCSAVDGLGLQQGLERLYEMILKRKKAARGGKKRR";
int charge = 15;
double intensityMultiplier = 1e6;
Proteomics.AminoAcidPolymer.Peptide baseSequence = new Proteomics.AminoAcidPolymer.Peptide(sequence);
var formula = baseSequence.GetChemicalFormula();
var isotopicDistribution = IsotopicDistribution.GetDistribution(formula, 0.125, 1e-8);
double[] masses = isotopicDistribution.Masses.ToArray();
double[] abundances = isotopicDistribution.Intensities.ToArray();
double max = abundances.Max();
List <(double, double)> peaks = new List <(double, double)>();
for (int i = 0; i < masses.Length; i++)
{
abundances[i] /= max;
if (abundances[i] >= 0.05)
{
peaks.Add((masses[i].ToMz(charge), abundances[i] * intensityMultiplier));
}
}
Random r = new Random(1);
for (int i = 0; i < 1000; i++)
{
double mz = r.NextDouble() * 1200 + 400;
double intensity = r.NextDouble() * 30000 + 30000;
peaks.Add((mz, intensity));
}
peaks = peaks.OrderBy(p => p.Item1).ToList();
var spectrum = new MzSpectrum(peaks.Select(p => p.Item1).ToArray(), peaks.Select(p => p.Item2).ToArray(), true);
var engine = new DeconvolutionEngine(0, 0.4, 3, 0.4, 1.5, 5, 1, 60, 2);
var envs = engine.Deconvolute(spectrum, spectrum.Range).ToList();
List <string> output = new List <string> {
DeconvolutedEnvelope.TabDelimitedHeader
};
foreach (var env in envs)
{
env.SpectraFileName = "temp";
output.Add(env.ToOutputString());
}
string path = Path.Combine(TestContext.CurrentContext.TestDirectory, @"TestPfmExplorerDeconOutput.tsv");
File.WriteAllLines(path, output);
var species = InputReaderParser.ReadSpeciesFromFile(path, out var errors);
Assert.That(!errors.Any());
Assert.That(species.Count > 0);
Assert.That(species.All(p => p.DeconvolutionFeature != null));
Assert.That(species.All(p => p.DeconvolutionFeature.AnnotatedEnvelopes != null && p.DeconvolutionFeature.AnnotatedEnvelopes.Count > 0));
Assert.That(species.All(p => p.DeconvolutionFeature.AnnotatedEnvelopes.All(v => v.PeakMzs.Count > 0)));
//Assert.That(species.Count == 1);
}
public static void TestCoIsolation()
{
Protease protease = new Protease("CustProtease", new List <Tuple <string, TerminusType> > {
new Tuple <string, TerminusType>("K", TerminusType.C)
}, new List <Tuple <string, TerminusType> >(), CleavageSpecificity.Full, null, null, null);
ProteaseDictionary.Dictionary.Add(protease.Name, protease);
CommonParameters CommonParameters = new CommonParameters(scoreCutoff: 1, deconvolutionIntensityRatio: 50, digestionParams: new DigestionParams(protease.Name, minPeptideLength: 1));
var variableModifications = new List <ModificationWithMass>();
var fixedModifications = new List <ModificationWithMass>();
var proteinList = new List <Protein> {
new Protein("MNNNKNDNK", null)
};
var searchModes = new SinglePpmAroundZeroSearchMode(5);
Proteomics.AminoAcidPolymer.Peptide pep1 = new Proteomics.AminoAcidPolymer.Peptide("NNNK");
Proteomics.AminoAcidPolymer.Peptide pep2 = new Proteomics.AminoAcidPolymer.Peptide("NDNK");
var dist1 = IsotopicDistribution.GetDistribution(pep1.GetChemicalFormula(), 0.1, 0.01);
var dist2 = IsotopicDistribution.GetDistribution(pep2.GetChemicalFormula(), 0.1, 0.01);
MsDataScan[] Scans = new MsDataScan[2];
double[] ms1intensities = new double[] { 0.8, 0.8, 0.2, 0.02, 0.2, 0.02 };
double[] ms1mzs = dist1.Masses.Concat(dist2.Masses).OrderBy(b => b).Select(b => b.ToMz(1)).ToArray();
double selectedIonMz = ms1mzs[1];
MzSpectrum MS1 = new MzSpectrum(ms1mzs, ms1intensities, false);
Scans[0] = new MsDataScan(MS1, 1, 1, false, Polarity.Positive, 1.0, new MzRange(300, 2000), "first spectrum", MZAnalyzerType.Unknown, MS1.SumOfAllY, null, null, "scan=1");
double[] ms2intensities = new double[] { 1, 1, 1, 1, 1 };
double[] ms2mzs = new double[] { 146.106.ToMz(1), 228.086.ToMz(1), 229.07.ToMz(1), 260.148.ToMz(1), 342.129.ToMz(1) };
MzSpectrum MS2 = new MzSpectrum(ms2mzs, ms2intensities, false);
double isolationMZ = selectedIonMz;
Scans[1] = new MsDataScan(MS2, 2, 2, false, Polarity.Positive, 2.0, new MzRange(100, 1500), "second spectrum", MZAnalyzerType.Unknown, MS2.SumOfAllY, null, null, "scan=2", selectedIonMz, null, null, isolationMZ, 2.5, DissociationType.HCD, 1, null);
var myMsDataFile = new MsDataFile(Scans, null);
bool DoPrecursorDeconvolution = true;
bool UseProvidedPrecursorInfo = true;
double DeconvolutionIntensityRatio = 50;
int DeconvolutionMaxAssumedChargeState = 10;
Tolerance DeconvolutionMassTolerance = new PpmTolerance(5);
var listOfSortedms2Scans = MetaMorpheusTask.GetMs2Scans(myMsDataFile, null, DoPrecursorDeconvolution, UseProvidedPrecursorInfo, DeconvolutionIntensityRatio, DeconvolutionMaxAssumedChargeState, DeconvolutionMassTolerance).OrderBy(b => b.PrecursorMass).ToArray();
PeptideSpectralMatch[] allPsmsArray = new PeptideSpectralMatch[listOfSortedms2Scans.Length];
List <ProductType> lp = new List <ProductType> {
ProductType.B, ProductType.Y
};
new ClassicSearchEngine(allPsmsArray, listOfSortedms2Scans, variableModifications, fixedModifications, proteinList, lp, searchModes, CommonParameters, new List <string>()).Run();
// Two matches for this single scan! Corresponding to two co-isolated masses
Assert.AreEqual(2, allPsmsArray.Length);
Assert.IsTrue(allPsmsArray[0].Score > 1);
Assert.AreEqual(2, allPsmsArray[0].ScanNumber);
var ojdfkj = (SequencesToActualProteinPeptidesEngineResults) new SequencesToActualProteinPeptidesEngine(new List <PeptideSpectralMatch>
{
allPsmsArray[0], allPsmsArray[1]
}, proteinList, fixedModifications, variableModifications, lp, new List <DigestionParams> {
CommonParameters.DigestionParams
}, CommonParameters.ReportAllAmbiguity, CommonParameters, new List <string>()).Run();
foreach (var huh in allPsmsArray)
{
if (huh != null)
{
huh.MatchToProteinLinkedPeptides(ojdfkj.CompactPeptideToProteinPeptideMatching);
}
}
Assert.AreEqual("NNNK", allPsmsArray[0].BaseSequence);
Assert.AreEqual("NDNK", allPsmsArray[1].BaseSequence);
}
public static void TestProteinQuantFileHeaders(bool hasDefinedExperimentalDesign, int bioreps, int fractions, int techreps)
{
// create the unit test directory
string unitTestFolder = Path.Combine(TestContext.CurrentContext.TestDirectory, @"TestProteinQuantFileHeaders");
Directory.CreateDirectory(unitTestFolder);
List <SpectraFileInfo> fileInfos = new List <SpectraFileInfo>();
string peptide = "PEPTIDE";
double ionIntensity = 1e6;
string condition = hasDefinedExperimentalDesign ? "TestCondition" : "";
// create the protein database
Protein prot = new Protein(peptide, @"");
string dbName = Path.Combine(unitTestFolder, "testDB.fasta");
UsefulProteomicsDatabases.ProteinDbWriter.WriteFastaDatabase(new List <Protein> {
prot
}, dbName, ">");
// create the .mzML files to search/quantify
for (int b = 0; b < bioreps; b++)
{
for (int f = 0; f < fractions; f++)
{
for (int r = 0; r < techreps; r++)
{
string fileToWrite = "file_" + "b" + b + "f" + f + "r" + r + ".mzML";
// generate mzml file
MsDataScan[] scans = new MsDataScan[2];
// create the MS1 scan
ChemicalFormula cf = new Proteomics.AminoAcidPolymer.Peptide(peptide).GetChemicalFormula();
IsotopicDistribution dist = IsotopicDistribution.GetDistribution(cf, 0.125, 1e-8);
double[] mz = dist.Masses.Select(v => v.ToMz(1)).ToArray();
double[] intensities = dist.Intensities.Select(v => v * ionIntensity).ToArray();
scans[0] = new MsDataScan(massSpectrum: new MzSpectrum(mz, intensities, false), oneBasedScanNumber: 1, msnOrder: 1, isCentroid: true,
polarity: Polarity.Positive, retentionTime: 1.0, scanWindowRange: new MzRange(400, 1600), scanFilter: "f",
mzAnalyzer: MZAnalyzerType.Orbitrap, totalIonCurrent: intensities.Sum(), injectionTime: 1.0, noiseData: null, nativeId: "scan=1");
// create the MS2 scan
var pep = new PeptideWithSetModifications(peptide, new Dictionary <string, Proteomics.Modification>());
List <Product> frags = new List <Product>();
pep.Fragment(DissociationType.HCD, FragmentationTerminus.Both, frags);
double[] mz2 = frags.Select(v => v.NeutralMass.ToMz(1)).ToArray();
double[] intensities2 = frags.Select(v => 1e6).ToArray();
scans[1] = new MsDataScan(massSpectrum: new MzSpectrum(mz2, intensities2, false), oneBasedScanNumber: 2, msnOrder: 2, isCentroid: true,
polarity: Polarity.Positive, retentionTime: 1.01, scanWindowRange: new MzRange(100, 1600), scanFilter: "f",
mzAnalyzer: MZAnalyzerType.Orbitrap, totalIonCurrent: intensities.Sum(), injectionTime: 1.0, noiseData: null, nativeId: "scan=2", selectedIonMz: pep.MonoisotopicMass.ToMz(1),
selectedIonChargeStateGuess: 1, selectedIonIntensity: 1e6, isolationMZ: pep.MonoisotopicMass.ToMz(1), isolationWidth: 1.5, dissociationType: DissociationType.HCD,
oneBasedPrecursorScanNumber: 1, selectedIonMonoisotopicGuessMz: pep.MonoisotopicMass.ToMz(1), hcdEnergy: "35");
// write the .mzML
string fullPath = Path.Combine(unitTestFolder, fileToWrite);
IO.MzML.MzmlMethods.CreateAndWriteMyMzmlWithCalibratedSpectra(
new MsDataFile(scans, new SourceFile(@"scan number only nativeID format", "mzML format", null, "SHA-1", @"C:\fake.mzML", null)),
fullPath, false);
var spectraFileInfo = new SpectraFileInfo(fullPath, condition, b, r, f);
fileInfos.Add(spectraFileInfo);
}
}
}
// write the experimental design for this quantification test
if (hasDefinedExperimentalDesign)
{
ExperimentalDesign.WriteExperimentalDesignToFile(fileInfos);
}
// run the search/quantification
SearchTask task = new SearchTask();
task.RunTask(unitTestFolder, new List <DbForTask> {
new DbForTask(dbName, false)
}, fileInfos.Select(p => p.FullFilePathWithExtension).ToList(), "");
// read in the protein quant results
Assert.That(File.Exists(Path.Combine(unitTestFolder, "AllQuantifiedProteinGroups.tsv")));
var lines = File.ReadAllLines(Path.Combine(unitTestFolder, "AllQuantifiedProteinGroups.tsv"));
// check the intensity column headers
var splitHeader = lines[0].Split(new char[] { '\t' }).ToList();
var intensityColumnHeaders = splitHeader.Where(p => p.Contains("Intensity", StringComparison.OrdinalIgnoreCase)).ToList();
Assert.That(intensityColumnHeaders.Count == 2);
if (!hasDefinedExperimentalDesign)
{
Assert.That(intensityColumnHeaders[0] == "Intensity_file_b0f0r0");
Assert.That(intensityColumnHeaders[1] == "Intensity_file_b1f0r0");
}
else
{
Assert.That(intensityColumnHeaders[0] == "Intensity_TestCondition_1");
Assert.That(intensityColumnHeaders[1] == "Intensity_TestCondition_2");
}
// check the protein intensity values
int ind1 = splitHeader.IndexOf(intensityColumnHeaders[0]);
int ind2 = splitHeader.IndexOf(intensityColumnHeaders[1]);
double intensity1 = double.Parse(lines[1].Split(new char[] { '\t' })[ind1]);
double intensity2 = double.Parse(lines[1].Split(new char[] { '\t' })[ind2]);
Assert.That(intensity1 > 0);
Assert.That(intensity2 > 0);
Assert.That(intensity1 == intensity2);
Directory.Delete(unitTestFolder, true);
}
public static void TestFlashLfqMatchBetweenRunsProteinQuant()
{
List <string> filesToWrite = new List <string> {
"mzml_1", "mzml_2"
};
List <string> pepSequences = new List <string> {
"PEPTIDE", "PEPTIDEV", "PEPTIDEVV", "PEPTIDEVVV", "PEPTIDEVVVV"
};
double intensity = 1e6;
double[] file1Rt = new double[] { 1.01, 1.02, 1.03, 1.04, 1.05 };
double[] file2Rt = new double[] { 1.015, 1.030, 1.036, 1.050, 1.065 };
Loaders.LoadElements(Path.Combine(TestContext.CurrentContext.TestDirectory, @"elements.dat"));
// generate mzml files (5 peptides each)
for (int f = 0; f < filesToWrite.Count; f++)
{
// 1 MS1 scan per peptide
MsDataScan[] scans = new MsDataScan[5];
for (int p = 0; p < pepSequences.Count; p++)
{
ChemicalFormula cf = new Proteomics.AminoAcidPolymer.Peptide(pepSequences[p]).GetChemicalFormula();
IsotopicDistribution dist = IsotopicDistribution.GetDistribution(cf, 0.125, 1e-8);
double[] mz = dist.Masses.Select(v => v.ToMz(1)).ToArray();
double[] intensities = dist.Intensities.Select(v => v * intensity).ToArray();
double rt;
if (f == 0)
{
rt = file1Rt[p];
}
else
{
rt = file2Rt[p];
}
// add the scan
scans[p] = new MsDataScan(massSpectrum: new MzSpectrum(mz, intensities, false), oneBasedScanNumber: p + 1, msnOrder: 1, isCentroid: true,
polarity: Polarity.Positive, retentionTime: rt, scanWindowRange: new MzRange(400, 1600), scanFilter: "f",
mzAnalyzer: MZAnalyzerType.Orbitrap, totalIonCurrent: intensities.Sum(), injectionTime: 1.0, noiseData: null, nativeId: "scan=" + (p + 1));
}
// write the .mzML
IO.MzML.MzmlMethods.CreateAndWriteMyMzmlWithCalibratedSpectra(new FakeMsDataFile(scans),
Path.Combine(TestContext.CurrentContext.TestDirectory, filesToWrite[f] + ".mzML"), false);
}
// set up spectra file info
SpectraFileInfo file1 = new SpectraFileInfo(Path.Combine(TestContext.CurrentContext.TestDirectory, filesToWrite[0] + ".mzML"), "a", 0, 0, 0);
SpectraFileInfo file2 = new SpectraFileInfo(Path.Combine(TestContext.CurrentContext.TestDirectory, filesToWrite[1] + ".mzML"), "a", 1, 0, 0);
// create some PSMs
var pg = new ProteinGroup("MyProtein", "gene", "org");
var myMbrProteinGroup = new ProteinGroup("MyMbrProtein", "MbrGene", "org");
Identification id1 = new Identification(file1, "PEPTIDE", "PEPTIDE",
new Proteomics.AminoAcidPolymer.Peptide("PEPTIDE").MonoisotopicMass, file1Rt[0] + 0.001, 1, new List <ProteinGroup> {
pg
});
Identification id2 = new Identification(file1, "PEPTIDEV", "PEPTIDEV",
new Proteomics.AminoAcidPolymer.Peptide("PEPTIDEV").MonoisotopicMass, file1Rt[1] + 0.001, 1, new List <ProteinGroup> {
pg
});
Identification id3 = new Identification(file1, "PEPTIDEVV", "PEPTIDEVV",
new Proteomics.AminoAcidPolymer.Peptide("PEPTIDEVV").MonoisotopicMass, file1Rt[2] + 0.001, 1, new List <ProteinGroup> {
myMbrProteinGroup
});
Identification id4 = new Identification(file1, "PEPTIDEVVV", "PEPTIDEVVV",
new Proteomics.AminoAcidPolymer.Peptide("PEPTIDEVVV").MonoisotopicMass, file1Rt[3] + 0.001, 1, new List <ProteinGroup> {
pg
});
Identification id5 = new Identification(file1, "PEPTIDEVVVV", "PEPTIDEVVVV",
new Proteomics.AminoAcidPolymer.Peptide("PEPTIDEVVVV").MonoisotopicMass, file1Rt[4] + 0.001, 1, new List <ProteinGroup> {
pg
});
Identification id6 = new Identification(file2, "PEPTIDE", "PEPTIDE",
new Proteomics.AminoAcidPolymer.Peptide("PEPTIDE").MonoisotopicMass, file2Rt[0] + 0.001, 1, new List <ProteinGroup> {
pg
});
Identification id7 = new Identification(file2, "PEPTIDEV", "PEPTIDEV",
new Proteomics.AminoAcidPolymer.Peptide("PEPTIDEV").MonoisotopicMass, file2Rt[1] + 0.001, 1, new List <ProteinGroup> {
pg
});
// missing ID 8 - MBR feature
Identification id9 = new Identification(file2, "PEPTIDEVVV", "PEPTIDEVVV",
new Proteomics.AminoAcidPolymer.Peptide("PEPTIDEVVV").MonoisotopicMass, file2Rt[3] + 0.001, 1, new List <ProteinGroup> {
pg
});
Identification id10 = new Identification(file2, "PEPTIDEVVVV", "PEPTIDEVVVV",
new Proteomics.AminoAcidPolymer.Peptide("PEPTIDEVVVV").MonoisotopicMass, file2Rt[4] + 0.001, 1, new List <ProteinGroup> {
pg
});
// test with top3 protein quant engine
FlashLFQEngine engine = new FlashLFQEngine(new List <Identification> {
id1, id2, id3, id4, id5, id6, id7, id9, id10
}, matchBetweenRuns: true);
var results = engine.Run();
Assert.That(results.ProteinGroups["MyMbrProtein"].GetIntensity(file1) > 0);
Assert.That(results.ProteinGroups["MyMbrProtein"].GetIntensity(file2) == 0);
// test with advanced protein quant engine
engine = new FlashLFQEngine(new List <Identification> {
id1, id2, id3, id4, id5, id6, id7, id9, id10
}, matchBetweenRuns: true, advancedProteinQuant: true);
results = engine.Run();
Assert.That(results.ProteinGroups["MyMbrProtein"].GetIntensity(file1) > 0);
Assert.That(results.ProteinGroups["MyMbrProtein"].GetIntensity(file2) == 0);
}
public static void TestFlashLfqAdvancedProteinQuant()
{
List <string> filesToWrite = new List <string> {
"mzml_1", "mzml_2"
};
List <string> pepSequences = new List <string> {
"PEPTIDE", "MYPEPTIDE", "VVVVVPEPTIDE"
};
double[,] amounts = new double[2, 3] {
{ 1000000, 1000000, 1000000 },
{ 2000000, 2000000, 900000 }
};
Loaders.LoadElements(Path.Combine(TestContext.CurrentContext.TestDirectory, @"elements.dat"));
// generate mzml files (3 peptides each)
for (int f = 0; f < filesToWrite.Count; f++)
{
// 1 MS1 scan per peptide
MsDataScan[] scans = new MsDataScan[3];
for (int p = 0; p < pepSequences.Count; p++)
{
ChemicalFormula cf = new Proteomics.AminoAcidPolymer.Peptide(pepSequences[p]).GetChemicalFormula();
IsotopicDistribution dist = IsotopicDistribution.GetDistribution(cf, 0.125, 1e-8);
double[] mz = dist.Masses.Select(v => v.ToMz(1)).ToArray();
double[] intensities = dist.Intensities.Select(v => v * amounts[f, p]).ToArray();
// add the scan
scans[p] = new MsDataScan(massSpectrum: new MzSpectrum(mz, intensities, false), oneBasedScanNumber: p + 1, msnOrder: 1, isCentroid: true,
polarity: Polarity.Positive, retentionTime: 1.0 + (p / 10.0), scanWindowRange: new MzRange(400, 1600), scanFilter: "f",
mzAnalyzer: MZAnalyzerType.Orbitrap, totalIonCurrent: intensities.Sum(), injectionTime: 1.0, noiseData: null, nativeId: "scan=" + (p + 1));
}
// write the .mzML
IO.MzML.MzmlMethods.CreateAndWriteMyMzmlWithCalibratedSpectra(new FakeMsDataFile(scans),
Path.Combine(TestContext.CurrentContext.TestDirectory, filesToWrite[f] + ".mzML"), false);
}
// set up spectra file info
SpectraFileInfo file1 = new SpectraFileInfo(Path.Combine(TestContext.CurrentContext.TestDirectory, filesToWrite[0] + ".mzML"), "a", 0, 0, 0);
SpectraFileInfo file2 = new SpectraFileInfo(Path.Combine(TestContext.CurrentContext.TestDirectory, filesToWrite[1] + ".mzML"), "a", 1, 0, 0);
// create some PSMs
var pg = new ProteinGroup("MyProtein", "gene", "org");
Identification id1 = new Identification(file1, "PEPTIDE", "PEPTIDE", 799.35996, 1.01, 1, new List <ProteinGroup> {
pg
});
Identification id2 = new Identification(file1, "MYPEPTIDE", "MYPEPTIDE", 1093.46377, 1.11, 1, new List <ProteinGroup> {
pg
});
Identification id3 = new Identification(file1, "VVVVVPEPTIDE", "VVVVVPEPTIDE", 1294.70203, 1.21, 1, new List <ProteinGroup> {
pg
});
Identification id4 = new Identification(file2, "PEPTIDE", "PEPTIDE", 799.35996, 1.01, 1, new List <ProteinGroup> {
pg
});
Identification id5 = new Identification(file2, "MYPEPTIDE", "MYPEPTIDE", 1093.46377, 1.11, 1, new List <ProteinGroup> {
pg
});
Identification id6 = new Identification(file2, "VVVVVPEPTIDE", "VVVVVPEPTIDE", 1294.70203, 1.21, 1, new List <ProteinGroup> {
pg
});
// create the FlashLFQ engine
FlashLFQEngine engine = new FlashLFQEngine(new List <Identification> {
id1, id2, id3, id4, id5, id6
}, normalize: false, advancedProteinQuant: true);
// run the engine
var results = engine.Run();
// third peptide should be low-weighted
// protein should be ~sum of first two peptide intensities (a little lower, because some smaller isotope peaks get skipped)
double file1ProteinIntensity = results.ProteinGroups["MyProtein"].GetIntensity(file1);
Assert.That(file1ProteinIntensity < 2e6);
Assert.That(file1ProteinIntensity > 1e6);
double file2ProteinIntensity = results.ProteinGroups["MyProtein"].GetIntensity(file2);
Assert.That(file2ProteinIntensity < 4e6);
Assert.That(file2ProteinIntensity > 3e6);
}
