In Mascot Server 2.3 and later, MS-MS searches can be run through Percolator. Percolator is produced by the MacCoss group at University of Washington. See e.g.
Percolator scores can be shown in Mascot Server reports instead of Mascot scores.
Percolated results are available both for old and new searches. However, they are only available when a decoy search has been performed. The Mascot results file is not changed by Percolator, but additional files for each results file are created by running
percolator.exe. The workflow is as follows:
DECOYoption is on.
ms-createpip.exeby calling setPercolatorFeatures() and getPercolatorFileNames().
ms-createpip.exe, specifying the output filename using the value from the step above.
percolator.exe, getting the required parameters by calling getPercolatorExeFlags() .
flags2parameter when creating an ms_peptidesummary object.
Steps 1 to 7 can be performed automatically after a search by specifying the appropriate options in getExecAfterSearch() .
There is also a static function, staticGetPercolatorFileNames() that can be called to get filenames without creating an
A 'Percolator score' is calculated from the posterior error probability (PEP) by
percolatorScore = -10 * log10(PEP)
Percolator processes only the rank 1 matches found by Mascot. Lower ranked peptides are scored by simply calculating
rank2PercolatorScore = (rank2MascotIonsScore/rank1MascotIonsScore) * rank1Percolatorscore
If the PEP value for rank 1 is exactly 1, it is reset to 0.9999. This is to ensure that there is a tiny amount of spread in scores for lower ranking peptides.
The function ms_peptide::getPercolatorScores() returns the posterior error probability which is used by Mascot Parser. The internal percolator score and the qValue can also be obtained, but these are unused by Mascot Parser.
When the MSPEPSUM_PERCOLATOR flag is specified, all Mascot scores are replaced with percolator derived scores. The original Mascot ions score for a peptide is still available by calling getPercolatorScores(). The following table describes the how each of the existing Mascot functions have been changed to return Percolator values.
|Mascot Parser Function||How value is calculated|
|ms_protein::getPeptideIonsScore()||Same value as getIonsScore() above, except a minor correction for large proteins is applied in the same way as for the Mascot Score.|
|ms_protein::getScore()||Calculated using the percolator scores rather than Mascot scores. Same rules for MudPIT and standard scoring apply.|
|ms_protein::getNonMudpitScore()||Calculated using the percolator scores rather than Mascot scores. Same rules for MudPIT scoring apply.|
|ms_mascotresults::getPeptideIdentityThreshold()||Calculated by taking |
|ms_mascotresults::getAvePeptideIdentityThreshold()||With Percolator, the threshold is the same for every query, so this is exactly the same as |
|ms_mascotresults::getMaxPeptideIdentityThreshold()||With Percolator, the threshold is the same for every query, so this is exactly the same as |
|ms_mascotresults::getHomologyThreshold()||With Percolator, there is no homology threshold, so this always returns 0.|
|ms_mascotresults::getHomologyThresholdForHistogram()||With Percolator, there is no homology threshold, so this always returns 0.|
|ms_mascotresults::getPeptideExpectationValue()||Return the posterior error probability by calculating it back from the score using |
|ms_mascotresults::getIonsScoreHistogram()||Returns a vector of Percolator scores rather than Mascot scores.|
|ms_mascotresults::getProteinScoreForHistogram()||Returns the Percolator protein score rather than Mascot protein score|
|ms_mascotresults::getNumHitsAboveIdentity()||Any peptide with a posterior error probability less than the significance value specified will be counted.|
|ms_mascotresults::getNumDecoyHitsAboveIdentity()||Any peptide with a posterior error probability less than the significance value specified will be counted.|
|ms_mascotresults::getNumHitsAboveHomology()||No homology thresholds, so always returns the same number as |
|ms_mascotresults::getNumDecoyHitsAboveHomology()||No homology thresholds, so always returns the same number as |
The following configuration functions are relevant:
|Copyright © 2016 Matrix Science Ltd. All Rights Reserved. Generated on Fri Jun 2 2017 01:44:51|