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MSGFPlus / msgfplus

MS-GF+ (aka MSGF+ or MSGFPlus) performs peptide identification by scoring MS/MS spectra against peptides derived from a protein sequence database.
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Help: ms-gf + stuck at 0% #64

Open GeorgesBed opened 5 years ago

GeorgesBed commented 5 years ago

Hello everyone, I've been using ms-gf+ for many years without any problem. However, recently on tmt data it's staying stuck at 0%.

below is a log example (running for 2 hours with 0% progress): MS-GF+ Release (v2019.04.18) (18 April 2019) Loading database files... Warning: Sequence database contains 240 counts of letter 'U', which does not correspond to an amino acid. Warning: Sequence database contains 17264 counts of letter 'X', which does not correspond to an amino acid. Loading database finished (elapsed time: 9.43 sec) Reading spectra... Ignoring 0 profile spectra. Ignoring 26 spectra having less than 10 peaks. Reading spectra finished (elapsed time: 25.53 sec) Using 24 threads. Search Parameters: PrecursorMassTolerance: 10.0 ppm IsotopeError: -1,2 TargetDecoyAnalysis: false FragmentationMethod: As written in the spectrum or CID if no info Instrument: QExactive (Q-Exactive) Enzyme: Tryp Protocol: TMT NumTolerableTermini: 2 MinPepLength: 6 MaxPepLength: 40 MinCharge: 2 MaxCharge: 5 NumMatchesPerSpec: 1 MaxMissedCleavages: -1 MaxNumModsPerPeptide: 3 ChargeCarrierMass: 1.00727649 (proton) MinNumPeaksPerSpectrum: 10 NumIsoforms: 128 Post translational modifications in use: Fixed (static): Carbamidomethyl on C (+57.0215) Fixed (static): TMT6plex on * at the peptide N-terminus (+229.1629) Fixed (static): TMT6plex on K (+229.1629) Variable (dynamic): Oxidation on M (+15.9949) Variable (dynamic): AcNoTMT on K (-187.1524)

Spectrum 0-12663 (total: 12664) Splitting work into 51 tasks. Search progress: 0 / 51 tasks, 0.00% 0.03 seconds elapsed Loading built-in param file: HCD_QExactive_Tryp_TMT.param Loading built-in param file: HCD_QExactive_Tryp_TMT.param Loading built-in param file: HCD_QExactive_Tryp_TMT.param Loading built-in param file: HCD_QExactive_Tryp_TMT.param Loading built-in param file: HCD_QExactive_Tryp_TMT.param Loading built-in param file: HCD_QExactive_Tryp_TMT.param Loading built-in param file: HCD_QExactive_Tryp_TMT.param Loading built-in param file: HCD_QExactive_Tryp_TMT.param Loading built-in param file: HCD_QExactive_Tryp_TMT.param Loading built-in param file: HCD_QExactive_Tryp_TMT.param Loading built-in param file: HCD_QExactive_Tryp_TMT.param Loading built-in param file: HCD_QExactive_Tryp_TMT.param Loading built-in param file: HCD_QExactive_Tryp_TMT.param Search progress: 0 / 51 tasks, 0.00% 1.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 2.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 3.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 4.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 5.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 6.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 7.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 8.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 9.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 10.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 11.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 12.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 13.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 14.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 15.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 16.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 17.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 18.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 19.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 20.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 21.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 22.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 23.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 24.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 25.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 26.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 27.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 28.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 29.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 30.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 31.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 32.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 33.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 34.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 35.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 36.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 37.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 38.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 39.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 40.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 41.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 42.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 43.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 44.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 45.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 46.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 47.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 48.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 49.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 50.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 51.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 52.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 53.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 54.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 55.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 56.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 57.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 58.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 59.00 minutes elapsed Search progress: 0 / 51 tasks, 0.00% 1.00 hours elapsed Search progress: 0 / 51 tasks, 0.00% 1.08 hours elapsed Search progress: 0 / 51 tasks, 0.00% 1.17 hours elapsed Search progress: 0 / 51 tasks, 0.00% 1.25 hours elapsed Search progress: 0 / 51 tasks, 0.00% 1.33 hours elapsed Search progress: 0 / 51 tasks, 0.00% 1.42 hours elapsed Search progress: 0 / 51 tasks, 0.00% 1.50 hours elapsed Search progress: 0 / 51 tasks, 0.00% 1.58 hours elapsed Search progress: 0 / 51 tasks, 0.00% 1.67 hours elapsed Search progress: 0 / 51 tasks, 0.00% 1.75 hours elapsed Search progress: 0 / 51 tasks, 0.00% 1.83 hours elapsed Search progress: 0 / 51 tasks, 0.00% 1.92 hours elapsed

Thank you.

GeorgesBed commented 5 years ago

Any toughts on how to fix this ?

alchemistmatt commented 5 years ago

You're using the latest release, so that's good. The log entries look normal. We haven't seen this issue, so the only way for us to diagnose would be if you send us your input files. Send me your e-mail address to proteomics@pnnl.gov and I can send you a link to use to transfer files to us. If you are unable to send us your data files in their entirety, you could use msconvert to convert to .mzML and export a subset of spectra (around 1000 spectra) and send us that.

GeorgesBed commented 5 years ago

I sent the file via wetransfer to proteomics@pnnl.gov Thank you for your help :)

alchemistmatt commented 5 years ago

I successfully retrieved the file. Testing with a 1.5 MB FASTA file ... I see the progress percentage increasing as expected. Thus, you'll also need to send me your FASTA file.

alchemistmatt commented 5 years ago

Also, are you using Windows or Linux? And what is your version of Java?
Use java -version On my Windows system I have:

java version "1.8.0_171"
Java(TM) SE Runtime Environment (build 1.8.0_171-b11)
Java HotSpot(TM) 64-Bit Server VM (build 25.171-b11, mixed mode)
alchemistmatt commented 5 years ago

171 is an old release; I updated to release 1.8.0_211, and that's working fine too. I'll next try JDK 12 from https://jdk.java.net/12/

alchemistmatt commented 5 years ago

Java 12 works fine too (though there are some warnings that I've documented at https://github.com/MSGFPlus/msgfplus/issues/26). Thus, the next step will be to test with your exact FASTA file.

GeorgesBed commented 5 years ago

I am using linux and the following version of java:

java version "1.8.0_60" Java(TM) SE Runtime Environment (build 1.8.0_60-b27) Java HotSpot(TM) 64-Bit Server VM (build 25.60-b23, mixed mode)

Surprisingly, I relaunched it and it is working just fine now even with the same database! It's worthy to mention that out of 200 mzML files only 19 had this issue. So it could be something hard to reproduce.

FarmGeek4Life commented 5 years ago

Are you running multiple searches at the same time? If you are searching the same mzML file with different databases at the same time, you could be encountering a file locking conflict (where one may complete, but another one never starts because of read/write locks on the file). I'm not sure on this, but it is a possibility.

There is another possible issue if multiple searches start at the same time for the same database - if the index files are not created, there can be conflicts with their creation (but this doesn't appear to be the problem).

GeorgesBed commented 5 years ago

Indeed, I am running on a cluster and the 200 searches were launched in parallel. but in my case, I'm using only one database (already indexed). So each mzML is used once and there should not be an index generation conflict.

alchemistmatt commented 5 years ago

Given that this is a transient error, it definitely looks like some sort of I/O issue. Glad to hear that it's fixed on a retry. When you launch the 200 searches, do you use any sort of delay between the launch of each search? If there are shared files, you might consider delaying the start of each search by 10, 15, or even 30 seconds.