I sent the link to your email. Thank you very much! On Wednesday, October 5, 2016 at 9:21:04 PM UTC-4, David Shteynberg wrote: > > I cannot tell from the search parameters if there is a problem. The only > issue I saw was that you didn't enable isotopic offsets, which are > sometimes helpful. If you send me a sample mzML file and possibly the > search results in pep.xml along with the database I can troubleshoot > further. One question, you said you are selecting 2+ 3+ and 4+, why are > only 2+ and 3+ in this dataset? Does any other dataset in your analysis > contain 4+? > > -David > > On Wed, Oct 5, 2016 at 6:16 PM, Ali <sma.ban...@gmail.com <javascript:>> > wrote: > >> Hi David >> >> We are only collecting 2+, 3+ and 4+ data. I am working on the cluster, >> so I am using the GPM version and we were unable to install the k-score >> plugin therefore I am using the X! Tandem native scoring for the searches. >> >> If you mean combining all of the search results with xinteract and then >> using PeptideProphet (as you instructed), I have not yet done so because I >> wanted to make sure that we are getting results from the individual >> searches. Is there an issue in my search parameters which is responsible >> for the problem? >> >> Many Thanks, >> Ali >> >> On Wednesday, October 5, 2016 at 7:06:19 PM UTC-4, David Shteynberg wrote: >>> >>> Hi Ali, >>> >>> >>> Are you setting your instrument to collect only 2+ and 3+ data? Are you >>> using the tandem bundled with TPP or the GPM version of tandem? Have you >>> tried to process all your runs in one file? >>> >>> Thanks, >>> -David >>> >>> >>> On Wed, Oct 5, 2016 at 11:39 AM, Ali <sma.ban...@gmail.com> wrote: >>> >>>> Thank you very much! I really appreciate it! >>>> >>>> On Tuesday, October 4, 2016 at 8:03:27 PM UTC-4, David Shteynberg wrote: >>>>> >>>>> PeptideProphet is doing it's job here by telling you that there is no >>>>> distinct population of correct results in this search. I suspect your >>>>> search parameters are the problem. This is orbitrap data so the charges >>>>> should be known, but it seems Tandem only searched 2+ and 3+ for you, >>>>> this >>>>> is somewhat troubling. Tomorrow, I will check your parameters more >>>>> carefully to try to identify the actual problem with your search. >>>>> >>>>> -David >>>>> >>>>> >>>>> On Tue, Oct 4, 2016 at 2:12 PM, Ali <sma.ban...@gmail.com> wrote: >>>>> >>>>>> It appears that PeptideProphet fails, I am getting this message: >>>>>> >>>>>> using Accurate Mass Bins >>>>>> using PPM mass difference >>>>>> (X! Tandem) (minprob 0) >>>>>> adding Accurate Mass mixture distr >>>>>> init with X! Tandem trypsin >>>>>> MS Instrument info: Manufacturer: Thermo Scientific, Model: UNKNOWN, >>>>>> Ionization: nanoelectrospray, Analyzer: orbitrap, Detector: inductive >>>>>> detector >>>>>> >>>>>> PeptideProphet (TPP v4.8.0 PHILAE, Build 201608051257-exported >>>>>> (RedHatEnterpriseServer-x86_64)) AKeller@ISB >>>>>> read in 0 1+, 14418 2+, 11080 3+, 0 4+, 0 5+, 0 6+, and 0 7+ spectra. >>>>>> Initialising statistical models ... >>>>>> negmean = 0.0533258 >>>>>> negmean = 1 >>>>>> negmean = 0.0376944 >>>>>> negmean = -0.2 >>>>>> negmean = -0.2 >>>>>> negmean = -0.2 >>>>>> negmean = -0.2 >>>>>> Iterations: .........10.........20. >>>>>> WARNING: Mixture model quality test failed for charge (2+). >>>>>> WARNING: Mixture model quality test failed for charge (3+). >>>>>> >>>>>> On Monday, October 3, 2016 at 5:57:39 PM UTC-4, David Shteynberg >>>>>> wrote: >>>>>>> >>>>>>> I highly recommend you start employing decoys in your search >>>>>>> databases. I noticed you are not combining together the runs in the >>>>>>> analysis. If the data was generated and searched in the same way it >>>>>>> should >>>>>>> be analysed together for better statistical power. I suggest you >>>>>>> combine >>>>>>> the results from all your runs and filter by the spectrum names later. >>>>>>> Try >>>>>>> this: >>>>>>> >>>>>>> InteractParser ./ALL.interact.xml ./*.tandem.pep.xml -S -Etrypsin >>>>>>> PeptideProphetParser ./ALL.interact.xml ACCMASS PPM NOICAT MINPROB=0 >>>>>>> >>>>>>> >>>>>>> -David >>>>>>> >>>>>>> On Mon, Oct 3, 2016 at 2:39 PM, Ali <sma.ban...@gmail.com> wrote: >>>>>>> >>>>>>>> Hi David >>>>>>>> >>>>>>>> Thanks for your reply. I don't think so, it is not just one set, as >>>>>>>> I said almost a third of my search results are returned with zero >>>>>>>> probabilities. All of the searches are done with the same setting and >>>>>>>> using >>>>>>>> the same database. The fact that for some of the searched files I am >>>>>>>> getting zero probabilities and some of them are just fine is a bit >>>>>>>> strange. >>>>>>>> I don't have decoys in my data base. If it helps, I can send you one >>>>>>>> of the >>>>>>>> search results with or without PeptideProphet probabilities. >>>>>>>> >>>>>>>> Sincerely, >>>>>>>> Ali >>>>>>>> >>>>>>>> >>>>>>>> >>>>>>>> On Monday, October 3, 2016 at 4:19:05 PM UTC-4, David Shteynberg >>>>>>>> wrote: >>>>>>>>> >>>>>>>>> Is it possible there are no correct results in that set? Common >>>>>>>>> pitfalls are incorrect search parameters (e.g. wrong mods enabled or >>>>>>>>> disabled) and wrong database. If you have decoys in your database >>>>>>>>> these >>>>>>>>> can be utilized for a more accurate mixture model generation using >>>>>>>>> options >>>>>>>>> "DECOY=<your decoy prefix> DECOYPROBS NONPARAM" in additions to the >>>>>>>>> options you already have enabled. >>>>>>>>> >>>>>>>>> -David >>>>>>>>> >>>>>>>>> On Mon, Oct 3, 2016 at 10:39 AM, Ali <sma.ban...@gmail.com> wrote: >>>>>>>>> >>>>>>>>>> Hi >>>>>>>>>> >>>>>>>>>> I am using PeptideProphet on my X!Tandem search results. There >>>>>>>>>> are ~150 experiments and PeptideProphet works fine on most of them >>>>>>>>>> but for >>>>>>>>>> ~50 experiments, PeptideProphet returns zero probability for all of >>>>>>>>>> the >>>>>>>>>> peptides. Can some one please tell me what might be the issue? >>>>>>>>>> >>>>>>>>>> *More Info:* >>>>>>>>>> I am using TPP v4.8 on Linux. PeptideProphet does not show an >>>>>>>>>> error when it produces zero probability for all of the search >>>>>>>>>> results. >>>>>>>>>> This is what I'm doing: >>>>>>>>>> -search with X!Tandem >>>>>>>>>> -Tandem2XML ./100.tandem.xml ./100.tandem.pep.xml >>>>>>>>>> -InteractParser ./100.interact.xml ./100.tandem.pep.xml -S >>>>>>>>>> -Etrypsin >>>>>>>>>> -PeptideProphetParser ./100.interact.xml ACCMASS PPM NOICAT >>>>>>>>>> MINPROB=0 >>>>>>>>>> >>>>>>>>>> >>>>>>>>>> Thank you very much, >>>>>>>>>> *Ali* >>>>>>>>>> >>>>>>>>>> -- >>>>>>>>>> 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 spctools-discu...@googlegroups.com. >>>>>>>>>> To post to this group, send email to spctools...@googlegroups.com >>>>>>>>>> . >>>>>>>>>> Visit this group at >>>>>>>>>> https://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. >>>>>>>> To unsubscribe from this group and stop receiving emails from it, >>>>>>>> send an email to spctools-discu...@googlegroups.com. >>>>>>>> To post to this group, send email to spctools...@googlegroups.com. >>>>>>>> Visit this group at >>>>>>>> https://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. >>>>>> To unsubscribe from this group and stop receiving emails from it, >>>>>> send an email to spctools-discu...@googlegroups.com. >>>>>> To post to this group, send email to spctools...@googlegroups.com. >>>>>> Visit this group at https://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. >>>> To unsubscribe from this group and stop receiving emails from it, send >>>> an email to spctools-discu...@googlegroups.com. >>>> To post to this group, send email to spctools...@googlegroups.com. >>>> Visit this group at https://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. >> To unsubscribe from this group and stop receiving emails from it, send an >> email to spctools-discu...@googlegroups.com <javascript:>. >> To post to this group, send email to spctools...@googlegroups.com >> <javascript:>. >> Visit this group at https://groups.google.com/group/spctools-discuss. >> For more options, visit https://groups.google.com/d/optout. >> > >
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