Hi David, I tried MINPROB=0.00, but I was still not able to get any peptides. It does not make sense to me, because my sample has plenty of peptides and the raw data do contain a lot of good MS2 spectra. In my opinion, the only reason is too many bad MS2 spectra included in the raw data.
My question is what settings should I use in X!Tandem and PeptideProphet/ProteinProphet to get the good MS2 spectra identified. Thanks. Xianyin. On Monday, March 10, 2014 2:19:04 PM UTC-4, David Shteynberg wrote: > All of your peptides have probabilities of 0 and since the setting shows > MINPROB=0.05, > all of the spetra are filtered from the resulting interact.pep.xml file. > This message is very telling "Found 11360 Decoys, and 10614 Non-Decoys", > assuming your decoy rate is 0.5 (rate of decoys amongst random matches) > then you have 0 correct results in this search. > > -David > > > On Mon, Mar 10, 2014 at 10:41 AM, <[email protected] <javascript:>>wrote: > >> One of my data failed and the return code 65280. The details are copied >> below. >> >> I noticed that I have more decoys than non-decoys. However, I found many >> good MS2 spectra when I manually check my raw data. >> >> What settings should I use to get the real good >> peptides validated through TPP (I used semi- when I did the database search >> using X!Tandem)? >> >> Thanks. >> >> Xianyin. >> >> >> ------------------------------------------------------------------------------------------------------------------------------------------------------- >> c:\Inetpub\tpp-bin\xinteract (TPP v4.6 OCCUPY rev 3, Build 201307241109 >> (MinGW)) >> PPM mode in Accurate Mass Model ... >> running: "C:/Inetpub/tpp-bin/InteractParser "interact.pep.xml" >> "20140307_12A.tandem.pep.xml" -L"7"" >> file 1: 20140307_12A.tandem.pep.xml >> SUCCESS: CORRECTED data file >> c:/Inetpub/wwwroot/ISB/data/20140307_12A.mzXML in msms_run_summary tag ... >> processed altogether 22005 results >> INFO: Results written to file: >> c:/Inetpub/wwwroot/ISB/data/interact.pep.xml >> command completed in 6 sec >> running: "C:/Inetpub/tpp-bin/DatabaseParser "interact.pep.xml"" >> command completed in 1 sec >> running: "C:/Inetpub/tpp-bin/RefreshParser "interact.pep.xml" >> "c:/Inetpub/wwwroot/ISB/data/RAT20140219T_DECOY.fasta"" >> - Searching the tree... >> - Linking duplicate entries... - Printing results... >> - Building Commentz-Walter keyword tree...command completed in 9 sec >> running: "C:/Inetpub/tpp-bin/PeptideProphetParser "interact.pep.xml" >> MINPROB=0.05 PPM ACCMASS DECOY=DECOY" >> using Accurate Mass Bins >> using PPM mass difference >> Using Decoy Label "DECOY". >> (X! Tandem (k-score)) >> adding Accurate Mass mixture distr >> init with X! Tandem (k-score) trypsin >> MS Instrument info: Manufacturer: Thermo Scientific, Model: LTQ Orbitrap >> Velos, Ionization: electrospray ionization, Analyzer: radial ejection >> linear ion trap, Detector: electron multiplier >> PeptideProphet (TPP v4.6 OCCUPY rev 3, Build 201307241109 (MinGW)) >> AKe...@ISB <javascript:> >> read in 0 1+, 6945 2+, 6638 3+, 4461 4+, 3930 5+, 0 6+, and 0 7+ spectra. >> Initialising statistical models ... >> Found 11360 Decoys, and 10614 Non-Decoys >> Iterations: .........10.........20.........30.. >> WARNING: Mixture model quality test failed for charge (2+). >> WARNING: Mixture model quality test failed for charge (3+). >> WARNING: Mixture model quality test failed for charge (4+). >> WARNING: Mixture model quality test failed for charge (5+). >> model complete after 33 iterations >> command completed in 34 sec >> running: "C:/Inetpub/tpp-bin/ProphetModels.pl -i interact.pep.xml -d >> "DECOY"" >> Analyzing interact.pep.xml ... >> Reading Accurate Mass Model model +1 ... >> Reading Accurate Mass Model model +2 ... >> Reading Accurate Mass Model model +3 ... >> Reading Accurate Mass Model model +4 ... >> Reading Accurate Mass Model model +5 ... >> Reading Accurate Mass Model model +6 ... >> Reading Accurate Mass Model model +7 ... >> Parsing search results "c:/Inetpub/wwwroot/ISB/data/20140307_12A (X! >> Tandem (k-score))"... >> => Found 0 hits. (0 decoys, 0 excluded) >> => Total so far: 0 hits. (0 decoys, 0 excluded) >> The system cannot find the path specified. >> command completed in 1 sec >> running: "c:/Inetpub/wwwroot/../tpp-bin/PepXMLViewer.cgi -I >> c:/Inetpub/wwwroot/ISB/data/interact.pep.xml" >> command "c:/Inetpub/wwwroot/../tpp-bin/PepXMLViewer.cgi -I >> c:/Inetpub/wwwroot/ISB/data/interact.pep.xml" failed: Unknown error >> command "c:/Inetpub/wwwroot/../tpp-bin/PepXMLViewer.cgi -I >> c:/Inetpub/wwwroot/ISB/data/interact.pep.xml" exited with non-zero exit >> code: 255 >> QUIT - the job is incomplete >> command "c:\Inetpub\tpp-bin\xinteract -Ninteract.pep.xml -p0.05 -l7 -PPM >> -OAp -dDECOY 20140307_12A.tandem.pep.xml" failed: Unknown error >> >> -- >> You received this message because you are subscribed to the Google Groups >> "spctools-discuss" group. >> To unsubscribe from this group and stop receiving emails from it, send an >> email to [email protected] <javascript:>. >> To post to this group, send email to >> [email protected]<javascript:> >> . >> Visit this group at http://groups.google.com/group/spctools-discuss. >> For more options, visit https://groups.google.com/d/optout. >> > > -- You received this message because you are subscribed to the Google Groups "spctools-discuss" group. 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