public void TestGetAllIsotopePeaks()
{
var methodName = MethodBase.GetCurrentMethod().Name;
TestUtils.ShowStarting(methodName);
const string specFilePath = @"H:\Research\GlycoTopDown\raw\User_sample_test_02252015.raw";
if (!File.Exists(specFilePath))
{
Assert.Ignore(@"Skipping test {0} since file not found: {1}", methodName, specFilePath);
}
//const int scanNum = 17338;
const double relativeIntensity = 0.1;
var run = PbfLcMsRun.GetLcMsRun(specFilePath);
var spec = run.GetSpectrum(17338);
var comp = Composition.Parse("C(610) H(945) N(172) O(189) S(3)");
var ion = new Ion(comp + BaseIonType.B.OffsetComposition, 9); // b127(9+)
Console.WriteLine("Composition: " + comp + " " + comp.Mass);
Console.WriteLine("b127(9+): " + ion.GetMonoIsotopicMz());
Console.WriteLine("b127(9+) 0th isotope: " + ion.GetIsotopeMz(0));
Console.WriteLine("b127(9+) 6th isotope: " + ion.GetIsotopeMz(6));
var peaks = spec.GetAllIsotopePeaks(ion, new Tolerance(10), relativeIntensity);
var isotopes = ion.GetIsotopes(relativeIntensity).ToArray();
for (var i = 0; i < isotopes.Length; i++)
{
if (peaks[i] == null) continue;
var isotopeIndex = isotopes[i].Index;
Console.WriteLine("{0}\t{1}\t{2}\t{3}", isotopeIndex, peaks[isotopeIndex].Mz, ion.GetIsotopeMz(isotopeIndex), GetPeakPpmError(peaks[isotopeIndex], ion.GetIsotopeMz(isotopeIndex)));
}
}
public void OutputStatistics(ProductSpectrum spectrum, Sequence sequence)
{
var baseIonTypes = spectrum.ActivationMethod != ActivationMethod.ETD ? BaseIonTypesCid : BaseIonTypesEtd;
var cleavages = sequence.GetInternalCleavages().ToArray();
var tolerance = new Tolerance(10);
var maxIntensity = spectrum.Peaks.Max(p => p.Intensity);
foreach (var c in cleavages)
{
foreach (var baseIonType in baseIonTypes)
{
var fragmentComposition = baseIonType.IsPrefix
? c.PrefixComposition + baseIonType.OffsetComposition
: c.SuffixComposition + baseIonType.OffsetComposition;
for (int charge = MinCharge; charge <= MaxCharge; charge++)
{
var ion = new Ion(fragmentComposition, charge);
var observedPeaks = spectrum.GetAllIsotopePeaks(ion, tolerance, RelativeIsotopeIntensityThreshold);
if (observedPeaks == null) continue;
var mostAbundantIsotopeIndex = ion.Composition.GetMostAbundantIsotopeZeroBasedIndex();
// representative peak intensity
var ionPeakIntensity = observedPeaks[mostAbundantIsotopeIndex].Intensity;
// calc. correlation
var isotopomerEnvelope = ion.Composition.GetIsotopomerEnvelopeRelativeIntensities();
var observedIntensities = new double[observedPeaks.Length];
for (var i = 0; i < observedPeaks.Length; i++)
{
var observedPeak = observedPeaks[i];
observedIntensities[i] = observedPeak != null ? (float)observedPeak.Intensity : 0.0;
}
var corrCoeff = FitScoreCalculator.GetPearsonCorrelation(isotopomerEnvelope, observedIntensities);
// mz error
var mostAbundantIsotopeMz = ion.GetIsotopeMz(mostAbundantIsotopeIndex);
var errorPpm = ((observedPeaks[mostAbundantIsotopeIndex].Mz - mostAbundantIsotopeMz)/
mostAbundantIsotopeMz)*1e6;
}
}
}
}
public void TestFitScoreCalculationCid()
{
var methodName = MethodBase.GetCurrentMethod().Name;
TestUtils.ShowStarting(methodName);
if (!File.Exists(TestLcMsRun.TestTopDownRawFilePathCid))
{
Assert.Ignore(@"Skipping test " + methodName + @" since file not found: " + TestLcMsRun.TestTopDownRawFilePathCid);
}
var run = InMemoryLcMsRun.GetLcMsRunScanRange(TestLcMsRun.TestTopDownRawFilePathCid, 5743, 5743);
var spec = run.GetSpectrum(5743);
Assert.True(spec != null);
const string protein = "MRIILLGAPGAGKGTQAQFIMEKYGIPQISTGDMLRAAVKSGSELGKQAKDIMDAGKLVTDELVIALVKERIAQEDCRNGFLLDGFPRTIPQADAMKEAGIVVDYVLEFDVPDELIVDRIVGRRVHAASGRVYHVKFNPPKVEGKDDVTGEDLTTRKDDQEETVRKRLVEYHQMTAPLIGYYQKEAEAGNTKYAKVDGTQAVADVRAALEKILG";
var protComp = new AminoAcidSet().GetComposition(protein) + Composition.H2O;
Assert.True(protComp != null);
Assert.True(protComp.C == 1035);
Assert.True(protComp.H == 1683);
Assert.True(protComp.N == 289);
Assert.True(protComp.O == 318);
Assert.True(protComp.P == 0);
Assert.True(protComp.S == 7);
Assert.True(Math.Abs(protComp.Mass - 23473.245267145) < 0.0000001);
Assert.True(protComp.NominalMass == 23461);
var ion = new Ion(protComp, 20);
// ion.Composition.ComputeApproximateIsotopomerEnvelop();
var isotopomerEnvelop = ion.Composition.GetIsotopomerEnvelopeRelativeIntensities();
Console.WriteLine(@"MonoMz: {0}, MonoMass: {1}", ion.GetMonoIsotopicMz(), ion.Composition.Mass);
var matchedPeaks = spec.GetAllIsotopePeaks(ion, new Tolerance(15), 0.1);
for (var i = 0; i < matchedPeaks.Length; i++)
{
Console.WriteLine(@"{0} {1} {2} {3}", i, ion.GetIsotopeMz(i), isotopomerEnvelop[i], matchedPeaks[i] == null ? 0 : matchedPeaks[i].Intensity);
}
var fitScore = spec.GetFitScore(ion, new Tolerance(15), 0.1);
var cosine = spec.GetConsineScore(ion, new Tolerance(15), 0.1);
var corr = spec.GetCorrScore(ion, new Tolerance(15), 0.1);
Console.WriteLine(@"FitScore: {0}", fitScore);
Console.WriteLine(@"Cosine: {0}", cosine);
Console.WriteLine(@"Corr: {0}", corr);
Assert.True(Math.Abs(fitScore - 0.181194589537041) < 0.0001);
Assert.True(Math.Abs(cosine - 0.917609346566222) < 0.0001);
Assert.True(Math.Abs(corr - 0.808326778009839) < 0.0001);
}
public void TestTopDownScoring()
{
var methodName = MethodBase.GetCurrentMethod().Name;
TestUtils.ShowStarting(methodName);
TopDownScorer.MaxCharge = 25;
TopDownScorer.MinCharge = 8;
const string specFilePath = @"C:\workspace\TopDown\E_coli_iscU_60_mock.raw";
const string protAnnotation = "A.AHAHLTHQYPAANAQVTAAPQAITLNFSEGVETGFSGAKITGPKNENIKTLPAKRNEQDQKQLIVPLADSLKPGTYTVDWHVVSVDGHKTKGHYTFSVK.";
var dehydro = new SearchModification(Modification.PyroGluQ, 'C', SequenceLocation.Everywhere, false);
var cysteinylC = new SearchModification(Modification.Cysteinyl, 'C', SequenceLocation.Everywhere, false);
var glutathioneC = new SearchModification(Modification.Glutathione, 'C', SequenceLocation.Everywhere, false);
var searchModifications = new List<SearchModification>
{
//pyroGluQ,
dehydro,
cysteinylC,
glutathioneC,
//oxM
};
//var aaSet = new AminoAcidSet(Modification.Carbamidomethylation);
var aaSet = new AminoAcidSet(searchModifications, 0);
var precursorTolerance = new Tolerance(10);
//Console.WriteLine(aaSet.GetAminoAcid('C').GetComposition());
// Create a sequence graph
//var protSeq = protAnnotation.Substring(2, protAnnotation.Length - 4);
var seqGraph = SequenceGraph.CreateGraph(aaSet, protAnnotation);
// TopDownScorer.MaxCharge = 60;
// TopDownScorer.MinCharge = 3;
var run = InMemoryLcMsRun.GetLcMsRun(specFilePath);
foreach (var protComposition in seqGraph.GetSequenceCompositions())
{
var mostAbundantIsotopeIndex = protComposition.GetMostAbundantIsotopeZeroBasedIndex();
Console.WriteLine("Composition\t{0}", protComposition);
Console.WriteLine("MostAbundantIsotopeIndex\t{0}", mostAbundantIsotopeIndex);
Console.WriteLine(new Ion(protComposition + Composition.H2O, 11).GetIsotopeMz(mostAbundantIsotopeIndex));
Console.WriteLine();
//for (var charge = TopDownScorer.MinCharge; charge <= TopDownScorer.MaxCharge; charge++)
//{
var scorer = new TopDownScorer(protComposition, run, precursorTolerance, null);
var score = scorer.GetScore();
Console.WriteLine(score);
//var precursorIon = new Ion(protComposition + Composition.H2O, charge);
//var xic = run.GetExtractedIonChromatogram(precursorIon.GetIsotopeMz(mostAbundantIsotopeIndex), precursorTolerance);
//Console.WriteLine(xic[0].ScanNum + " " + xic[1].ScanNum);
//Console.WriteLine("ScanNum\t{0}", string.Join("\t", xic.Select(p => p.ScanNum.ToString())));
//Console.WriteLine("precursorCharge " + charge + "\t" + string.Join("\t", xic.Select(p => p.Intensity.ToString())));
// }
Console.WriteLine("\nCharge\tm/z");
for (var charge = 9; charge <= 18; charge++)
{
var precursorIon = new Ion(protComposition + Composition.H2O, charge);
Console.WriteLine("{0}\t{1}", charge, precursorIon.GetIsotopeMz(mostAbundantIsotopeIndex));
}
}
// sw.Stop();
// Console.WriteLine(@"Elapsed Time: {0:f4} sec", sw.Elapsed.TotalSeconds);
}
private double[][][] GetXicArray()
{
var scanNumberIndex = new Dictionary<int, int>();
var i = 0;
foreach (var scanNumber in _run.GetScanNumbers(1))
{
// Console.WriteLine(scanNumber + " " + i);
scanNumberIndex[scanNumber] = i++;
}
var xicArray = new double[MaxCharge - MinCharge + 1][][];
// Console.WriteLine("***");
for (var charge = MinCharge; charge <= MaxCharge; charge++)
{
var xicArrayForThisCharge = new double[_isotopeEnvelope.Length][];
for (var j = 0; j < xicArrayForThisCharge.Length; j++)
xicArrayForThisCharge[j] = new double[scanNumberIndex.Count];
var precursorIon = new Ion(_proteinCompositionPlusWater, charge);
// Console.Write("precursorCharge " + charge);
for (var k = 0; k < xicArrayForThisCharge.Length; k++)
{
var mz = precursorIon.GetIsotopeMz(k + _minIsotopeIndex);
var xic = _run.GetPrecursorExtractedIonChromatogram(mz, _tolerance);
//Console.WriteLine("Mz" + mz);
foreach (var xicPeak in xic)
{
// Console.WriteLine(xicPeak.MostAbundantIsotopeMz);
xicArrayForThisCharge[k][scanNumberIndex[xicPeak.ScanNum]] = xicPeak.Intensity;
// if (k == _maxIntensityIsotopeIndex-_minIsotopeIndex)
// Console.WriteLine(charge + "\t" + xicPeak.MostAbundantIsotopeMz + "\t" + xicPeak.Intensity + "\t" + scanNumberIndex[xicPeak.MostAbundantIsotopeMz]);
//if(k == 0 && charge == 13) Console.WriteLine(xicPeak.MostAbundantIsotopeMz + " " + xicPeak.Intensity);
}
// Console.WriteLine();
}
// System.Environment.Exit(1);
//
xicArray[charge - MinCharge] = xicArrayForThisCharge;
}
return xicArray;
}
private bool FindIon(Ion ion, Tolerance tolerance, double relativeIntensityThreshold, out int baseIsotopePeakIndex, out int nIsotopes, out int nMatchedIsotopes)
{
//matchedPeakIndex = new List<int>();
var baseIsotopeIndex = ion.Composition.GetMostAbundantIsotopeZeroBasedIndex();
var isotopomerEnvelope = ion.Composition.GetIsotopomerEnvelopeRelativeIntensities();
var baseIsotopMz = ion.GetIsotopeMz(baseIsotopeIndex);
baseIsotopePeakIndex = _ms2Spec.FindPeakIndex(baseIsotopMz, tolerance);
nIsotopes = isotopomerEnvelope.Select(x => x >= relativeIntensityThreshold).Count();
nMatchedIsotopes = 0;
if (baseIsotopePeakIndex < 0) return false;
//if (baseIsotopePeakIndex < 0) baseIsotopePeakIndex = ~baseIsotopePeakIndex;
nMatchedIsotopes++;
// go down
var peakIndex = baseIsotopePeakIndex;
//matchedPeakIndex.Add(peakIndex);
for (var isotopeIndex = baseIsotopeIndex - 1; isotopeIndex >= 0; isotopeIndex--)
{
if (isotopomerEnvelope[isotopeIndex] < relativeIntensityThreshold) break;
var isotopeMz = ion.GetIsotopeMz(isotopeIndex);
var tolTh = tolerance.GetToleranceAsTh(isotopeMz);
var minMz = isotopeMz - tolTh;
var maxMz = isotopeMz + tolTh;
for (var i = peakIndex - 1; i >= 0; i--)
{
var peakMz = _ms2Spec.Peaks[i].Mz;
if (peakMz < minMz)
{
//peakIndex = i;
//break;
return false;
}
if (peakMz <= maxMz) // find match, move to prev isotope
{
peakIndex = i;
//matchedPeakIndex.Add(peakIndex);
nMatchedIsotopes++;
break;
}
}
}
// go up
peakIndex = baseIsotopePeakIndex;
for (var isotopeIndex = baseIsotopeIndex + 1; isotopeIndex < isotopomerEnvelope.Length; isotopeIndex++)
{
if (isotopomerEnvelope[isotopeIndex] < relativeIntensityThreshold) break;
var isotopeMz = ion.GetIsotopeMz(isotopeIndex);
var tolTh = tolerance.GetToleranceAsTh(isotopeMz);
var minMz = isotopeMz - tolTh;
var maxMz = isotopeMz + tolTh;
for (var i = peakIndex + 1; i < _ms2Spec.Peaks.Length; i++)
{
var peakMz = _ms2Spec.Peaks[i].Mz;
if (peakMz > maxMz)
{
//peakIndex = i;
//break;
return false;
}
if (peakMz >= minMz) // find match, move to prev isotope
{
peakIndex = i;
//matchedPeakIndex.Add(peakIndex);
nMatchedIsotopes++;
break;
}
}
}
return true;
}
public void AnalyizeFusionDdaData()
{
var methodName = MethodBase.GetCurrentMethod().Name;
TestUtils.ShowStarting(methodName);
// Parameters
//const double relativeIntensityThreshold = 0.7;
const double precursorTolerancePpm = 20;
const string specFilePath = @"D:\Research\Data\UW\Fusion\WT_D_DDA_130412065618.raw";
var run = InMemoryLcMsRun.GetLcMsRun(specFilePath);
const double fdrThreshold = 0.01;
var tolerance = new Tolerance(precursorTolerancePpm);
var aaSet = new AminoAcidSet(Modification.Carbamidomethylation);
const string resultFilePath = @"D:\Research\Data\UW\Fusion\WT_D_DDA_130412065618_10ppm_TI2_SGD_Decoy.tsv";
Console.WriteLine("IsDecoy\tPeptide\tScanNum\tCharge\tSpecEValue\tQValue\tPrecursorMz" +
"\tTheo0\tTheo1\tTheo2\tTheo3" +
"\tObs0\tCorr0\tObs1\tCorr1\tObs2\tCorr2\tObs3\tCorr3\tObs-1\tCorr-1\tObs0.5\tCorr0.5");
foreach (var line in File.ReadLines(resultFilePath))
{
if (line.StartsWith("#")) continue;
var token = line.Split('\t');
if (token.Length != 16) continue;
var qValue = Convert.ToDouble(token[14]);
if (qValue > fdrThreshold) continue;
var peptide = token[8].Replace("C+57.021", "C");
var scanNum = Convert.ToInt32(token[2]);
var charge = Convert.ToInt32(token[7]);
var specEValue = Convert.ToDouble(token[12]);
var protein = token[9];
var isDecoy = protein.StartsWith("XXX_");
var precursorIon = new Ion(aaSet.GetComposition(peptide) + Composition.H2O, charge);
var baseXic = run.GetPrecursorExtractedIonChromatogram(precursorIon.GetMostAbundantIsotopeMz(), tolerance, scanNum);
var baseIntensity = baseXic.GetSumIntensities();
Console.Write("{0}\t{1}\t{2}\t{3}\t{4}\t{5}\t{6}", (isDecoy ? 1 : 0), peptide, scanNum, charge, specEValue, qValue, precursorIon.GetMonoIsotopicMz());
var isotopeIndices = new double[] {0, 1, 2, 3, -1, 0.5};
var theoIsotopes = precursorIon.GetIsotopes(0.01);
var numIsotopes = 0;
foreach (var theoIsotope in theoIsotopes)
{
Console.Write("\t"+theoIsotope.Ratio);
if (++numIsotopes == 4) break;
}
foreach (var isotopeIndex in isotopeIndices)
{
var isotopeMz = precursorIon.GetIsotopeMz(isotopeIndex);
var xic = run.GetPrecursorExtractedIonChromatogram(isotopeMz, tolerance, scanNum);
var relativeIntensity = xic.GetSumIntensities() / baseIntensity;
var correlation = xic.GetCorrelation(baseXic);
Console.Write("\t{0}\t{1}", relativeIntensity, correlation);
}
Console.WriteLine();
}
}
/// <summary>
/// Checks whether this spectrum contains all isotope peaks whose relative intensity is equal or larter than the threshold
/// </summary>
/// <param name="ion">ion</param>
/// <param name="tolerance">tolerance</param>
/// <param name="relativeIntensityThreshold">relative intensity threshold of the theoretical isotope profile</param>
/// <returns>true if spectrum contains all ions; false otherwise.</returns>
public bool ContainsIon(Ion ion, Tolerance tolerance, double relativeIntensityThreshold)
{
var baseIsotopeIndex = ion.Composition.GetMostAbundantIsotopeZeroBasedIndex();
var isotopomerEnvelope = ion.Composition.GetIsotopomerEnvelopeRelativeIntensities();
var baseIsotopMz = ion.GetIsotopeMz(baseIsotopeIndex);
var baseIsotopePeakIndex = FindPeakIndex(baseIsotopMz, tolerance);
if (baseIsotopePeakIndex < 0) return false;
// go down
var peakIndex = baseIsotopePeakIndex;
for (var isotopeIndex = baseIsotopeIndex - 1; isotopeIndex >= 0; isotopeIndex--)
{
if (isotopomerEnvelope[isotopeIndex] < relativeIntensityThreshold) break;
var isotopeMz = ion.GetIsotopeMz(isotopeIndex);
var tolTh = tolerance.GetToleranceAsTh(isotopeMz);
var minMz = isotopeMz - tolTh;
var maxMz = isotopeMz + tolTh;
for (var i = peakIndex - 1; i >= 0; i--)
{
var peakMz = Peaks[i].Mz;
if (peakMz < minMz) return false;
if (peakMz <= maxMz) // find match, move to prev isotope
{
peakIndex = i;
break;
}
}
}
// go up
peakIndex = baseIsotopePeakIndex;
for (var isotopeIndex = baseIsotopeIndex + 1; isotopeIndex < isotopomerEnvelope.Length; isotopeIndex++)
{
if (isotopomerEnvelope[isotopeIndex] < relativeIntensityThreshold) break;
var isotopeMz = ion.GetIsotopeMz(isotopeIndex);
var tolTh = tolerance.GetToleranceAsTh(isotopeMz);
var minMz = isotopeMz - tolTh;
var maxMz = isotopeMz + tolTh;
for (var i = peakIndex + 1; i < Peaks.Length; i++)
{
var peakMz = Peaks[i].Mz;
if (peakMz > maxMz) return false;
if (peakMz >= minMz) // find match, move to prev isotope
{
peakIndex = i;
break;
}
}
}
return true;
}
public void TestXicGen()
{
var methodName = MethodBase.GetCurrentMethod().Name;
TestUtils.ShowStarting(methodName);
const string specFilePath = @"D:\Research\Data\UW\Fusion\WT_D_DDA_130412065618.raw";
var run = InMemoryLcMsRun.GetLcMsRun(specFilePath);
// Test
var tolerance = new Tolerance(30);
const string peptide = "AIANGQVDGFPTQEECR";
const int targetScanNum = 37633;
const int charge = 2;
//const string peptide = "IVDTNGAGDAFAGGFMAGLTK";
//const int targetScanNum = 67513;
//const int charge = 3;
var aaSet = new AminoAcidSet(Modification.Carbamidomethylation);
var precursorIon = new Ion(aaSet.GetComposition(peptide) + Composition.H2O, charge);
Console.WriteLine("Theoretical isotopomer profile:");
foreach(var p in precursorIon.GetIsotopes(0.1)) Console.WriteLine("{0}\t{1}", precursorIon.GetIsotopeMz(p.Index), p.Ratio);
var xicArr = new Dictionary<int, Xic>();
var basePeakIndex = precursorIon.Composition.GetMostAbundantIsotopeZeroBasedIndex();
for (var i = -1; i < 3; i++)
{
xicArr[i] = run.GetPrecursorExtractedIonChromatogram(precursorIon.GetIsotopeMz(i), tolerance, targetScanNum);
}
for (var i = -1; i < 3; i++)
{
Console.WriteLine("\nIndex: {0}", i);
Console.WriteLine("m/z: {0}", precursorIon.GetIsotopeMz(i));
Console.WriteLine("#XicPeaks: {0}", xicArr[i].Count);
Console.WriteLine("Intensity: {0}", xicArr[i].GetSumIntensities()/xicArr[basePeakIndex].GetSumIntensities());
Console.WriteLine("Correlation: {0}", xicArr[i].GetCorrelation(xicArr[basePeakIndex]));
}
}
public void TestIsotopemerProfileByKyowon() // is faster and more accurate than IsotopicDistributionCalculator
{
var methodName = MethodBase.GetCurrentMethod().Name;
ShowStarting(methodName);
//C78H120N22O28S3 C150H120N220O28S30
//var additionalElements = new[]
// {
// new Tuple<Atom, short>(Atom.Get("P"), 1),
// new Tuple<Atom, short>(Atom.Get("13C"), 3),
// new Tuple<Atom, short>(Atom.Get("15N"), 1),
// };
//var composition = new Composition(149, 244, 44, 57, 0, additionalElements);
// var composition = new Composition(83, 136, 22, 24, 1);
//var composition = new Composition(210, 323, 54, 61, 0);
var composition = new Composition(419, 699, 119, 129, 1);
const int charge = 14;
var ion = new Ion(composition + Composition.H2O, charge);
var ff = composition.GetIsotopomerEnvelopeRelativeIntensities();
var isotopeIndex = -1;
foreach (var ii in ff)
{
++isotopeIndex;
Console.WriteLine("{0}: {1}\t{2}", isotopeIndex, ion.GetIsotopeMz(isotopeIndex), ii);
}
}
public void TestIsoProfile()
{
var methodName = MethodBase.GetCurrentMethod().Name;
ShowStarting(methodName);
var composition = Composition.Parse("C(82) H(149) N(23) O(24) S(3)");
const int charge = 3;
var ion = new Ion(composition, charge);
foreach (var isotope in ion.GetIsotopes(0.1))
{
Console.WriteLine("{0}\t{1}\t{2}", isotope.Index, ion.GetIsotopeMz(isotope.Index), isotope.Ratio);
}
Console.WriteLine(composition.Mass);
}
public void TestPeptide()
{
var methodName = MethodBase.GetCurrentMethod().Name;
ShowStarting(methodName);
//const string sequence = "MSGRGKGGKGLGKGGAKRHRKVLRDNIQGITKPAIRRLARRGGVKRISGLIYEETRGVLKVFLENVIRDAVTYTEHAKRKTVTAMDVVYALKRQGRTLYGFGG"; // Histone H4
const string sequence = "IRDAVTYTEHAKRKTVTAMDVVYALKRQGRTLYGFGG"; // Histone H4
//const string sequence = "MRIILLGAPGAGKGTQAQFIMEKYGIPQISTGDMLRAAVKSGSELGKQAKDIMDAGKLVTDELVIALVKERIAQEDCRNGFLLDGFPRTIPQADAMKEAGIVVDYVLEFDVPDELIVDRIVGRRVHAASGRVYHVKFNPPKVEGKDDVTGEDLTTRKDDQEETVRKRLVEYHQMTAPLIGYYQKEAEAGNTKYAKVDGTQAVADVRAALEKILG";
//const string sequence = "MNKTQLIDVIAEKAELSKTQAKAALESTLAAITESLKEGDAVQLVGFGTFKVNHRAERTGRNPQTGKEIKIAAANVPAFVSGKALKDAVK";
//const string sequence =
// "METTKPSFQDVLEFVRLFRRKNKLQREIQDVEKKIRDNQKRVLLLDNLSDYIKPGMSVEAIQGIIASMKGDYEDRVDDYIIKNAELSKERRDISKKLKAMGEMKNGEAK";
var aaSet = new AminoAcidSet();
var composition = aaSet.GetComposition(sequence) + Composition.H2O;
Console.WriteLine(composition);
Console.WriteLine(composition.Mass);
Console.WriteLine(composition.NominalMass);
// 2nd isotope
Console.WriteLine(composition.GetIsotopeMass(0));
Console.WriteLine(composition.GetIsotopeMass(1));
Console.WriteLine(composition.GetIsotopeMass(2));
//Assert.AreEqual(composition.ToPlainString(), "C34H51N7O14");
Console.WriteLine("Isotopomer Envelope:");
foreach (var e in composition.GetIsotopomerEnvelopeRelativeIntensities()) Console.WriteLine(e);
Console.WriteLine();
Console.WriteLine("Isotope ions:");
var ion = new Ion(composition + Composition.H2O, 13);
foreach (var p in ion.GetIsotopes(0.1)) Console.WriteLine("{0}\t{1}", ion.GetIsotopeMz(p.Index), p.Ratio);
Console.WriteLine();
}
public void TestIsoProfile()
{
var methodName = MethodBase.GetCurrentMethod().Name;
TestUtils.ShowStarting(methodName);
const string sequence = "MWYMISAQDVENSLEKRLAARPAHLARLQELADEGRLLVAGPHPAIDSENPGDAGFSGSLVVADFDSLATAQAWADADPYFAAGVYQSVVVKPFKRVLP";
var aaSet = new AminoAcidSet();
var comp = aaSet.GetComposition(sequence) + Composition.H2O;
var ion = new Ion(comp, 9);
foreach (var i in ion.GetIsotopes(0.1))
{
Console.WriteLine(ion.GetIsotopeMz(i.Index)+"\t"+i.Ratio);
}
}
public void TestGeneratingXicsOfAllCharges()
{
var methodName = MethodBase.GetCurrentMethod().Name;
TestUtils.ShowStarting(methodName);
if (!File.Exists(TestRawFilePath))
{
Assert.Ignore(@"Skipping test " + methodName + @" since file not found: " + TestRawFilePath);
}
var run = PbfLcMsRun.GetLcMsRun(TestRawFilePath, 0.0, 0.0);
var comparer = new MzComparerWithBinning(27);
const string protSequence =
"AIPQSVEGQSIPSLAPMLERTTPAVVSVAVSGTHVSKQRVPDVFRYFFGPNAPQEQVQERPFRGLGSGVIIDADKGYIVTNNHVIDGADDIQVGLHDGREVKAKLIGTDSESDIALLQIEAKNLVAIKTSDSDELRVGDFAVAIGNPFGLGQTVTSGIVSALGRSGLGIEMLENFIQTDAAINSGNSGGALVNLKGELIGINTAIVAPNGGNVGIGFAIPANMVKNLIAQIAEHGEVRRGVLGIAGRDLDSQLAQGFGLDTQHGGFVNEVSAGSAAEKAGIKAGDIIVSVDGRAIKSFQELRAKVATMGAGAKVELGLIRDGDKKTVNVTLGEANQTTEKAAGAVHPMLQGASLENASKGVEITDVAQGSPAAMSGLQKGDLIVGINRTAVKDLKSLKELLKDQEGAVALKIVRGKSMLYLVLR";
//const string annotation = "_." + protSequence + "._";
var seqGraph = SequenceGraph.CreateGraph(new AminoAcidSet(), AminoAcid.ProteinNTerm, protSequence, AminoAcid.ProteinCTerm);
if (seqGraph == null) return;
seqGraph.SetSink(0);
var neutral = seqGraph.GetSinkSequenceCompositionWithH2O() - Composition.Hydrogen;
var proteinMass = neutral.Mass;
var isoEnv = Averagine.GetIsotopomerEnvelope(proteinMass);
const bool SHOW_ALL_SCANS = false;
var targetColIndex = 0;
#pragma warning disable 0162
if (SHOW_ALL_SCANS)
Console.WriteLine("Charge\t" + string.Join("\t", run.GetScanNumbers(1)));
else
{
// Just display data for scan 161
Console.WriteLine("Charge\t161");
foreach (var scanNumber in run.GetScanNumbers(1))
{
if (scanNumber == 161)
break;
targetColIndex++;
}
}
#pragma warning restore 0162
const int minCharge = 2;
const int maxCharge = 60;
for (var charge = minCharge; charge <= maxCharge; charge++)
{
var ion = new Ion(neutral, charge);
var mostAbundantIsotopeMz = ion.GetIsotopeMz(isoEnv.MostAbundantIsotopeIndex);
//var secondMostAbundantIsotopeMz = ion.GetIsotopeMz(isoEnv.MostAbundantIsotopeIndex + 1);
var binNum = comparer.GetBinNumber(mostAbundantIsotopeMz);
var mzStart = comparer.GetMzStart(binNum);
var mzEnd = comparer.GetMzEnd(binNum);
var xic = run.GetFullPrecursorIonExtractedIonChromatogram(mzStart, mzEnd);
Console.Write(charge+"\t");
#pragma warning disable 0162
if (SHOW_ALL_SCANS)
Console.WriteLine(string.Join("\t", xic.Select(p => p.Intensity)));
else
Console.WriteLine(xic[targetColIndex].Intensity);
#pragma warning restore 0162
}
}
public void GetIsoProfile()
{
var methodName = MethodBase.GetCurrentMethod().Name;
TestUtils.ShowStarting(methodName);
const string protSequence =
"AIPQSVEGQSIPSLAPMLERTTPAVVSVAVSGTHVSKQRVPDVFRYFFGPNAPQEQVQERPFRGLGSGVIIDADKGYIVTNNHVIDGADDIQVGLHDGREVKAKLIGTDSESDIALLQIEAKNLVAIKTSDSDELRVGDFAVAIGNPFGLGQTVTSGIVSALGRSGLGIEMLENFIQTDAAINSGNSGGALVNLKGELIGINTAIVAPNGGNVGIGFAIPANMVKNLIAQIAEHGEVRRGVLGIAGRDLDSQLAQGFGLDTQHGGFVNEVSAGSAAEKAGIKAGDIIVSVDGRAIKSFQELRAKVATMGAGAKVELGLIRDGDKKTVNVTLGEANQTTEKAAGAVHPMLQGASLENASKGVEITDVAQGSPAAMSGLQKGDLIVGINRTAVKDLKSLKELLKDQEGAVALKIVRGKSMLYLVLR";
//const string annotation = "_." + protSequence + "._";
var seqGraph = SequenceGraph.CreateGraph(new AminoAcidSet(), AminoAcid.ProteinNTerm, protSequence, AminoAcid.ProteinCTerm);
if (seqGraph == null) return;
seqGraph.SetSink(0);
var neutral = seqGraph.GetSinkSequenceCompositionWithH2O() - Composition.Hydrogen;
//Console.WriteLine(neutral);
for (var charge = 22; charge <= 60; charge++)
{
var ion = new Ion(neutral, charge);
Console.WriteLine("{0}\t{1}", charge, ion.GetMostAbundantIsotopeMz());
}
var ion27 = new Ion(neutral, 29);
var isotopes = ion27.GetIsotopes(0.1);
foreach (var isotope in isotopes)
{
Console.WriteLine("{0}\t{1}", ion27.GetIsotopeMz(isotope.Index), isotope.Ratio);
}
}
public Peak[] GetAllIsotopePeaks(Spectrum spec, Ion ion, Tolerance tolerance, double relativeIntensityThreshold, out int[] peakIndexList)
{
var mostAbundantIsotopeIndex = ion.Composition.GetMostAbundantIsotopeZeroBasedIndex();
var isotopomerEnvelope = ion.Composition.GetIsotopomerEnvelopeRelativeIntensities();
peakIndexList = new int[isotopomerEnvelope.Length];
var mostAbundantIsotopeMz = ion.GetIsotopeMz(mostAbundantIsotopeIndex);
var mostAbundantIsotopeMatchedPeakIndex = spec.FindPeakIndex(mostAbundantIsotopeMz, tolerance);
if (mostAbundantIsotopeMatchedPeakIndex < 0) return null;
var observedPeaks = new Peak[isotopomerEnvelope.Length];
observedPeaks[mostAbundantIsotopeIndex] = spec.Peaks[mostAbundantIsotopeMatchedPeakIndex];
peakIndexList[mostAbundantIsotopeIndex] = mostAbundantIsotopeMatchedPeakIndex;
// go down
var peakIndex = mostAbundantIsotopeMatchedPeakIndex - 1;
for (var isotopeIndex = mostAbundantIsotopeIndex - 1; isotopeIndex >= 0; isotopeIndex--)
{
if (isotopomerEnvelope[isotopeIndex] < relativeIntensityThreshold) break;
var isotopeMz = ion.GetIsotopeMz(isotopeIndex);
var tolTh = tolerance.GetToleranceAsTh(isotopeMz);
var minMz = isotopeMz - tolTh;
var maxMz = isotopeMz + tolTh;
for (var i = peakIndex; i >= 0; i--)
{
var peakMz = spec.Peaks[i].Mz;
if (peakMz < minMz)
{
peakIndex = i;
break;
}
if (peakMz <= maxMz) // find match, move to prev isotope
{
var peak = spec.Peaks[i];
if (observedPeaks[isotopeIndex] == null ||
peak.Intensity > observedPeaks[isotopeIndex].Intensity)
{
observedPeaks[isotopeIndex] = peak;
peakIndexList[isotopeIndex] = i;
}
}
}
}
// go up
peakIndex = mostAbundantIsotopeMatchedPeakIndex + 1;
for (var isotopeIndex = mostAbundantIsotopeIndex + 1; isotopeIndex < isotopomerEnvelope.Length; isotopeIndex++)
{
if (isotopomerEnvelope[isotopeIndex] < relativeIntensityThreshold) break;
var isotopeMz = ion.GetIsotopeMz(isotopeIndex);
var tolTh = tolerance.GetToleranceAsTh(isotopeMz);
var minMz = isotopeMz - tolTh;
var maxMz = isotopeMz + tolTh;
for (var i = peakIndex; i < spec.Peaks.Length; i++)
{
var peakMz = spec.Peaks[i].Mz;
if (peakMz > maxMz)
{
peakIndex = i;
break;
}
if (peakMz >= minMz) // find match, move to prev isotope
{
var peak = spec.Peaks[i];
if (observedPeaks[isotopeIndex] == null ||
peak.Intensity > observedPeaks[isotopeIndex].Intensity)
{
observedPeaks[isotopeIndex] = peak;
peakIndexList[isotopeIndex] = i;
}
}
}
}
return observedPeaks;
}
public void TestFusionDdaData()
{
var methodName = MethodBase.GetCurrentMethod().Name;
TestUtils.ShowStarting(methodName);
// Parameters
const double relativeIntensityThreshold = 0.7;
const double precursorTolerancePpm = 20;
//const double isotopeRatioTolerance = 2;
//const double correlationThreshold = 0.3;
const double fdrThreshold = 0.01;
const string specFilePath = @"D:\Research\Data\UW\Fusion\WT_D_DDA_130412065618.raw";
var run = InMemoryLcMsRun.GetLcMsRun(specFilePath);
var sw = new System.Diagnostics.Stopwatch();
sw.Start();
var tolerance = new Tolerance(precursorTolerancePpm);
var aaSet = new AminoAcidSet(Modification.Carbamidomethylation);
const string resultFilePath = @"D:\Research\Data\UW\Fusion\oldResult\WT_D_DDA_130412065618_10ppm_TI2_SGD_Decoy.tsv";
var numTargets = 0;
var numValidTargets = 0;
var numDecoys = 0;
var numValidDecoys = 0;
foreach(var line in File.ReadLines(resultFilePath))
{
if (line.StartsWith("#")) continue;
var token = line.Split('\t');
if (token.Length != 16) continue;
var qValue = Convert.ToDouble(token[14]);
if (qValue > fdrThreshold) continue;
var peptide = token[8].Replace("C+57.021", "C");
var scanNum = Convert.ToInt32(token[2]);
var charge = Convert.ToInt32(token[7]);
var protein = token[9];
var isDecoy = protein.StartsWith("XXX_");
if (isDecoy) numDecoys++;
else numTargets++;
var precursorIon = new Ion(aaSet.GetComposition(peptide) + Composition.H2O, charge);
var basePeakIndex = precursorIon.Composition.GetMostAbundantIsotopeZeroBasedIndex();
var baseXic = run.GetPrecursorExtractedIonChromatogram(precursorIon.GetMostAbundantIsotopeMz(), tolerance, scanNum);
var baseIntensity = baseXic.GetSumIntensities();
var isValid = true;
foreach (var isotope in precursorIon.GetIsotopes(relativeIntensityThreshold))
{
if (isotope.Index == basePeakIndex) continue;
var isotopeMz = precursorIon.GetIsotopeMz(isotope.Index);
var xic = run.GetPrecursorExtractedIonChromatogram(isotopeMz, tolerance, scanNum);
if (xic.Count == 0)
{
isValid = false;
break;
}
//if (xic.Count > 0)
//{
// var isotopeRatio = xic.GetSumIntensities() / baseIntensity / isotope.Item2;
// var correlation = xic.GetCorrelation(baseXic);
// if (isotopeRatio > 0.8 && isotopeRatio < 1.2
// && correlation > 0.8)
// {
// isValid = true;
// }
//}
// Check if isotope ratio is within tolerance
//if (isotopeRatio > isotopeRatioTolerance || isotopeRatio < 1 / isotopeRatioTolerance)
//{
// isValid = false;
// //Console.WriteLine("Off ratio\t{0}\t{1}\t{2}\t{3}\t{4}\t{5}\t{6}", isDecoy, peptide, scanNum, charge, precursorIon.GetMonoIsotopicMz(), isotopeMz, isotopeRatio);
// break;
//}
// Check if correlation is high
//if (correlation < correlationThreshold)
//{
// isValid = false;
// //Console.WriteLine("Low correlation\t{0}\t{1}\t{2}\t{3}\t{4}\t{5}\t{6}", isDecoy, peptide, scanNum, charge, precursorIon.GetMonoIsotopicMz(), isotopeMz, correlation);
// break;
//}
}
if (isValid && !isDecoy) numValidTargets++;
else if (isValid) numValidDecoys++;
//Console.WriteLine("{0}\t{1}\t{2}", peptide, scanNum, charge);
}
Console.WriteLine("#Targets: {0}", numTargets);
Console.WriteLine("#ValidTargets: {0}\t{1}", numValidTargets, numValidTargets/(double)numTargets);
Console.WriteLine("#Decoys: {0}", numDecoys);
Console.WriteLine("#ValidDecoys: {0}\t{1}", numValidDecoys, numValidDecoys / (double)numDecoys);
sw.Stop();
Console.WriteLine(@"TimeForPrecursorValidation {0:f4} sec", sw.Elapsed.TotalSeconds);
}
public void GetIntensity(string peptideSequence, double ppmTolerance)
{
Composition composition;
if (peptideSequence.Equals("Tetraoctylammonium"))
{
composition = new Composition(32, 67, 1, 0, 0);
}
else if (peptideSequence.Equals("Tetraoctylammonium Bromide"))
{
composition = new Composition(64, 135, 2, 0, 0) + Composition.ParseFromPlainString("Br");
}
else
{
composition = PeptideUtil.GetCompositionOfPeptide(peptideSequence);
}
string empiricalFormula = composition.ToPlainString();
for (int chargeState = 1; chargeState <= 5; chargeState++)
{
// Calculate Target m/z
var targetIon = new Ion(composition, chargeState);
double targetMz = targetIon.GetMonoIsotopicMz();
double minMzForSpectrum = targetMz - (3.0 / chargeState);
double maxMzForSpectrum = targetMz + (10.0 / chargeState);
Console.WriteLine(peptideSequence + " - +" + chargeState + " - " + targetMz);
// Generate Theoretical Isotopic Profile
IsotopicProfile theoreticalIsotopicProfile = _theoreticalFeatureGenerator.GenerateTheorProfile(empiricalFormula, chargeState);
List<Peak> theoreticalIsotopicProfilePeakList = theoreticalIsotopicProfile.Peaklist.Cast<Peak>().ToList();
// Find XIC Features
IEnumerable<FeatureBlob> featureBlobs = FindFeatures(targetMz, ppmTolerance, 1, 1);
// Check each XIC Peak found
foreach (var featureBlob in featureBlobs)
{
FeatureBlobStatistics statistics = featureBlob.CalculateStatistics();
int unsaturatedIsotope = 0;
FeatureBlob isotopeFeature = null;
int scanLcMin = statistics.ScanLcMin;
int scanLcMax = statistics.ScanLcMax;
int scanImsMin = statistics.ScanImsMin;
int scanImsMax = statistics.ScanImsMax;
double intensity = statistics.SumIntensities;
// Find an unsaturated peak in the isotopic profile
for (int i = 1; i < 10; i++)
{
if (!statistics.IsSaturated) break;
// Target isotope m/z
double isotopeTargetMz = targetIon.GetIsotopeMz(i);
// Find XIC Features
IEnumerable<FeatureBlob> newFeatureBlobs = FindFeatures(isotopeTargetMz, ppmTolerance, 1, 1);
// If no feature, then get out
if (!newFeatureBlobs.Any())
{
statistics = null;
break;
}
bool foundFeature = false;
foreach (var newFeatureBlob in newFeatureBlobs.OrderByDescending(x => x.PointList.Count))
{
var newStatistics = newFeatureBlob.CalculateStatistics();
if (newStatistics.ScanImsRep <= scanImsMax && newStatistics.ScanImsRep >= scanImsMin && newStatistics.ScanLcRep <= scanLcMax && newStatistics.ScanLcRep >= scanLcMin)
{
isotopeFeature = newFeatureBlob;
foundFeature = true;
break;
}
}
if (!foundFeature)
{
statistics = null;
break;
}
statistics = isotopeFeature.CalculateStatistics();
unsaturatedIsotope = i;
}
int scanImsRep = statistics.ScanImsRep;
// Get ViperCompatibleMass Spectrum Data
XYData massSpectrum = GetMassSpectrum(1, scanImsMin, scanImsMax, scanImsRep, minMzForSpectrum, maxMzForSpectrum);
//List<Peak> massSpectrumPeakList = _peakDetector.FindPeaks(massSpectrum);
//WriteXYDataToFile(massSpectrum, targetMz);
// Find Isotopic Profile
List<Peak> massSpectrumPeaks;
IsotopicProfile observedIsotopicProfile = _msFeatureFinder.IterativelyFindMSFeature(massSpectrum, theoreticalIsotopicProfile, out massSpectrumPeaks);
double unsaturatedIntensity = observedIsotopicProfile != null ? observedIsotopicProfile.GetSummedIntensity() : 0;
// Correct for Saturation if needed
if (unsaturatedIsotope > 0)
{
IsotopicProfileUtil.AdjustSaturatedIsotopicProfile(observedIsotopicProfile, theoreticalIsotopicProfile, unsaturatedIsotope);
}
if (observedIsotopicProfile != null && observedIsotopicProfile.MonoIsotopicMass > 1)
{
Console.WriteLine("ScanIMS = " + scanImsMin + "-" + scanImsMax + "\tImsRep = " + scanImsRep + "\tUncorrectedIntensity = " + unsaturatedIntensity + "\tIntensity = " + observedIsotopicProfile.GetSummedIntensity());
}
}
}
}