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compomics / peptide-shaker

Interpretation of proteomics identification results
http://compomics.github.io/projects/peptide-shaker.html
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Error when using Peptideshaker to open the files produced from SearchGUI #274

Closed joeyuzh closed 4 years ago

joeyuzh commented 7 years ago

Hi,

Recently, I have encountered an issue that Peptideshaker got stuck at "Validating identifications at 1% FDR" step when trying to process the files from SearchGUI. The report was attached below.

Wed Jul 12 09:21:11 PDT 2017 Import process for PPvsDR_concise (Sample: PPvsDR_concise_0711, Replicate: 0)

Wed Jul 12 09:21:11 PDT 2017 Importing sequences from uniprot-proteome%253AUP000002524_DR_concatenated_target_decoy.fasta. Wed Jul 12 09:21:13 PDT 2017 FASTA file import completed. Wed Jul 12 09:21:13 PDT 2017 Importing gene mappings. Wed Jul 12 09:21:13 PDT 2017 Deinococcus radiodurans (strain ATCC 13939 / DSM 20539 / JCM 16871 / LMG 4051 / NBRC 15346 / NCIMB 9279 / R1 / VKM B-1422) not available in Ensembl. Gene and GO annotation for this species will not be available. Wed Jul 12 09:21:14 PDT 2017 Establishing local database connection. Wed Jul 12 09:21:22 PDT 2017 Reading identification files. Wed Jul 12 09:21:22 PDT 2017 Parsing 20170512_BC_PP_0.msgf.mzid. Wed Jul 12 09:21:31 PDT 2017 Loading spectra for 20170512_BC_PP_0.msgf.mzid. Wed Jul 12 09:21:31 PDT 2017 Importing 20170512_BC_PP_0.mgf Wed Jul 12 09:21:31 PDT 2017 20170512_BC_PP_0.mgf imported. Wed Jul 12 09:21:31 PDT 2017 Collecting peptides to map. Wed Jul 12 09:21:32 PDT 2017 Mapping peptides to proteins. Wed Jul 12 09:21:32 PDT 2017 Importing PSMs from 20170512_BC_PP_0.msgf.mzid Wed Jul 12 09:21:57 PDT 2017 Parsing 20170512_BC_PP_0.t.xml. Wed Jul 12 09:22:16 PDT 2017 Loading spectra for 20170512_BC_PP_0.t.xml. Wed Jul 12 09:22:16 PDT 2017 Collecting peptides to map. Wed Jul 12 09:22:16 PDT 2017 Mapping peptides to proteins. Wed Jul 12 09:22:17 PDT 2017 Importing PSMs from 20170512_BC_PP_0.t.xml Wed Jul 12 09:22:18 PDT 2017 9715 identified spectra (24.4%) did not present a valid peptide. Wed Jul 12 09:22:18 PDT 2017 14137 of the best scoring peptides were excluded by the import filters: Wed Jul 12 09:22:18 PDT 2017 - 31.3% peptide length less than 8 or greater than 30. Wed Jul 12 09:22:18 PDT 2017 - 64.1% peptide presenting high mass or isotopic deviation. Wed Jul 12 09:22:18 PDT 2017 - 4.5% unrecognized modifications. Wed Jul 12 09:22:18 PDT 2017 File import completed. 82855 first hits imported (423505 secondary) from 52232 spectra. Wed Jul 12 09:22:18 PDT 2017 [68751 first hits passed the initial filtering] Wed Jul 12 09:22:18 PDT 2017 Computing assumptions probabilities. Wed Jul 12 09:22:18 PDT 2017 Saving assumptions probabilities. Wed Jul 12 09:22:19 PDT 2017 Selecting best peptide per spectrum. Wed Jul 12 09:22:30 PDT 2017 Computing PSM probabilities. Wed Jul 12 09:22:30 PDT 2017 Scoring PTMs in PSMs (D-score and PhosphoRS) Wed Jul 12 09:24:20 PDT 2017 Resolving peptide inference issues. Wed Jul 12 09:24:33 PDT 2017 Saving probabilities, building peptides and proteins. Wed Jul 12 09:28:12 PDT 2017 Simplifying protein groups. Wed Jul 12 09:28:12 PDT 2017 27 unlikely protein mappings found: Wed Jul 12 09:28:12 PDT 2017 - 19 protein groups supported by non-enzymatic shared peptides. Wed Jul 12 09:28:12 PDT 2017 - 14 groups explained by a simpler group. Wed Jul 12 09:28:12 PDT 2017 Note: a group can present combinations of these criteria. Wed Jul 12 09:28:12 PDT 2017 Generating peptide map. Wed Jul 12 09:30:02 PDT 2017 Computing peptide probabilities. Wed Jul 12 09:30:02 PDT 2017 Saving peptide probabilities. Wed Jul 12 09:33:42 PDT 2017 Generating protein map. Wed Jul 12 09:34:13 PDT 2017 Resolving protein inference issues, inferring peptide and protein PI status. Wed Jul 12 09:49:46 PDT 2017 60 conflicts resolved. 7 protein groups remaining (1 suspicious). Wed Jul 12 09:49:46 PDT 2017 Correcting protein probabilities. Wed Jul 12 09:49:46 PDT 2017 Saving protein probabilities. Wed Jul 12 09:52:00 PDT 2017 Validating identifications at 1% FDR, quality control of matches. Wed Jul 12 10:03:04 PDT 2017 An error occurred: null

hbarsnes commented 7 years ago

Hi Joe,

Thanks for telling us about this. Could you share your PeptideShaker log file with us? This should tell us more about why the error occurs. You'll find the file via the PeptideShaker Welcome dialog > Settings & Help > Help > Bug Report.

Best regards, Harald

joeyuzh commented 7 years ago

Hi Harald,

Thanks for your reply. The bug report can be seen below.

Wed Jul 12 09:19:15 PDT 2017: PeptideShaker version 1.16.11. Memory given to the Java virtual machine: 3817865216. Total amount of memory in the Java virtual machine: 128974848. Free memory: 82342320. Java version: 1.8.0_131. 1714 script command tokens (C) 2009 Jmol Development Jmol Version: 12.0.43 2011-05-03 14:21 java.vendor: Oracle Corporation java.version: 1.8.0_131 os.name: Windows 7 memory: 54.5/163.1 processors available: 8 useCommandThread: false java.lang.NullPointerException at eu.isas.peptideshaker.validation.MatchesValidator$PeptideValidatorRunnable.run(MatchesValidator.java:1668) at java.util.concurrent.Executors$RunnableAdapter.call(Unknown Source) at java.util.concurrent.FutureTask.run(Unknown Source) at java.util.concurrent.ThreadPoolExecutor.runWorker(Unknown Source) at java.util.concurrent.ThreadPoolExecutor$Worker.run(Unknown Source) at java.lang.Thread.run(Unknown Source)

hbarsnes commented 7 years ago

Hi Joe,

Thanks for the log. It seems like something is going wrong when validating one of your peptides. Would it be possible for you to share the data with us so that we can look deeper into the issue? We would need all the data you are trying to load in PeptideShaker.

Best regards, Harald

joeyuzh commented 7 years ago

Hi Harald,

Thanks so much for your help. The zip file generated from peptideshaker and the FASTA file are attached.

​ searchgui_out.zip https://drive.google.com/file/d/0B_kSNm8C402TdFZ4ekhtYWk4RGM/view?usp=drive_web ​ ​ uniprot-proteome%253AUP000002524_DR_concatenate... https://drive.google.com/file/d/0B_kSNm8C402TVmtPRFZSa19JTjA/view?usp=drive_web ​

Regards,

Joe

On Thu, Jul 13, 2017 at 8:58 AM, Harald Barsnes notifications@github.com wrote:

Hi Joe,

Thanks for the log. It seems like something is going wrong when validating one of your peptides. Would it be possible for you to share the data with us so that we can look deeper into the issue? We would need all the data you are trying to load in PeptideShaker.

Best regards, Harald

— You are receiving this because you authored the thread. Reply to this email directly, view it on GitHub https://github.com/compomics/peptide-shaker/issues/274#issuecomment-315122483, or mute the thread https://github.com/notifications/unsubscribe-auth/AJ8rV1LyKs3UOgRZoV298wGD_0Dqhu5Kks5sNj69gaJpZM4OWI3e .

hbarsnes commented 7 years ago

Hi Joe,

Thanks for the data. However, I will also need the mgf file to be able to load the data in PeptideShaker. Could you share this file as well?

Best regards, Harald

joeyuzh commented 7 years ago

Hi Harald,

I'm sorry that mgf was missed in the last email. It is attached below.

​ 20170512_BC_PP_0.mgf https://drive.google.com/file/d/0B_kSNm8C402TcnU2eGtpVXdiN0U/view?usp=drive_web ​

Warm regards,

Joe

On Thu, Jul 13, 2017 at 2:27 PM, Harald Barsnes notifications@github.com wrote:

Hi Joe,

Thanks for the data. However, I will also need the mgf file to be able to load the data in PeptideShaker. Could you share this file as well?

Best regards, Harald

— You are receiving this because you authored the thread. Reply to this email directly, view it on GitHub https://github.com/compomics/peptide-shaker/issues/274#issuecomment-315207022, or mute the thread https://github.com/notifications/unsubscribe-auth/AJ8rVxv_EVWpKv9qE_as7UL0MrXEhc24ks5sNou3gaJpZM4OWI3e .

hbarsnes commented 7 years ago

Hi Joe,

Thanks for the mgf file. I have now started to process your data in PeptideShaker. I'm already passed the "Validating identifications at 1% FDR, quality control of matches" step though and it all seems to work fine. Perhaps you could try simply reloading the data and see if that solves the problem?

BTW, is there anything special with regards to your computer setup? For example, are the data stored on a network drive or at a location with automatic backup that could interfere with the files during the analysis?

Best regards, Harald

joeyuzh commented 7 years ago

Hi Harald,

Thanks for your information. I will try it again to see if it works.

Regards,

Joe

On Fri, Jul 14, 2017 at 4:08 AM, Harald Barsnes notifications@github.com wrote:

Hi Joe,

Thanks for the mgf file. I have now started to process your data in PeptideShaker. I'm already passed the "Validating identifications at 1% FDR, quality control of matches" step though and it all seems to work fine. Perhaps you could try simply reloading the data and see if that solves the problem?

BTW, is there anything special with regards to your computer setup? For example, are the data stored on a network drive or at a location with automatic backup that could interfere with the files during the analysis?

Best regards, Harald

— You are receiving this because you authored the thread. Reply to this email directly, view it on GitHub https://github.com/compomics/peptide-shaker/issues/274#issuecomment-315334160, or mute the thread https://github.com/notifications/unsubscribe-auth/AJ8rV5LerExfrMz3pdF1WQ4iZ0Y-hpaPks5sN0wYgaJpZM4OWI3e .

joeyuzh commented 7 years ago

Hi Harald,

I have tried to run the file in another computer. However, it stilled failed at the "Validating identifications at 1% FDR, quality control of matches" step. The snapshot of the error message is attached below. I'm wondering which version of Peptideshaker do you use? The one installed on our lab's computer is 1.16.11. The OS is windows 7 and the computer's memory is 16 GB.

​

Regards,

Joe

On Fri, Jul 14, 2017 at 9:37 AM, Joe Yu joebonic00@gmail.com wrote:

Hi Harald,

Thanks for your information. I will try it again to see if it works.

Regards,

Joe

On Fri, Jul 14, 2017 at 4:08 AM, Harald Barsnes notifications@github.com wrote:

Hi Joe,

Thanks for the mgf file. I have now started to process your data in PeptideShaker. I'm already passed the "Validating identifications at 1% FDR, quality control of matches" step though and it all seems to work fine. Perhaps you could try simply reloading the data and see if that solves the problem?

BTW, is there anything special with regards to your computer setup? For example, are the data stored on a network drive or at a location with automatic backup that could interfere with the files during the analysis?

Best regards, Harald

— You are receiving this because you authored the thread. Reply to this email directly, view it on GitHub https://github.com/compomics/peptide-shaker/issues/274#issuecomment-315334160, or mute the thread https://github.com/notifications/unsubscribe-auth/AJ8rV5LerExfrMz3pdF1WQ4iZ0Y-hpaPks5sN0wYgaJpZM4OWI3e .

hbarsnes commented 7 years ago

Hi Joe,

I'm using the 1.16.11 version as well. Windows 10 and for this test I only gave PeptideShaker 4 GB of memory.

The snapshot of the error message is attached below.

The snapshot did not make it through the GitHub issue system I'm afraid. You have to upload them via the issue tracker, adding as attachment to the email does not work.

Best regards, Harald

joeyuzh commented 7 years ago

Hi Harald,

The complete error report of the file run on another computer is attached below. Thanks for your time and advice.

Regards,

Joe

Fri Jul 14 11:00:00 PDT 2017 Import process for PPvsDR_concise (Sample: PPvsDR_concise_0714, Replicate: 0)

Fri Jul 14 11:00:00 PDT 2017 Importing sequences from uniprot-proteome%253AUP000002524_DR_concatenated_target_decoy.fasta. Fri Jul 14 11:00:03 PDT 2017 FASTA file import completed. Fri Jul 14 11:00:03 PDT 2017 Importing gene mappings. Fri Jul 14 11:00:03 PDT 2017 Deinococcus radiodurans (strain ATCC 13939 / DSM 20539 / JCM 16871 / LMG 4051 / NBRC 15346 / NCIMB 9279 / R1 / VKM B-1422) not available in Ensembl. Gene and GO annotation for this species will not be available. Fri Jul 14 11:00:03 PDT 2017 Establishing local database connection. Fri Jul 14 11:00:11 PDT 2017 Reading identification files. Fri Jul 14 11:00:11 PDT 2017 Parsing 20170512_BC_PP_0.msgf.mzid. Fri Jul 14 11:00:24 PDT 2017 Loading spectra for 20170512_BC_PP_0.msgf.mzid. Fri Jul 14 11:00:24 PDT 2017 Importing 20170512_BC_PP_0.mgf Fri Jul 14 11:00:26 PDT 2017 20170512_BC_PP_0.mgf imported. Fri Jul 14 11:00:26 PDT 2017 Collecting peptides to map. Fri Jul 14 11:00:27 PDT 2017 Mapping peptides to proteins. Fri Jul 14 11:00:29 PDT 2017 Importing PSMs from 20170512_BC_PP_0.msgf.mzid Fri Jul 14 11:00:37 PDT 2017 Parsing 20170512_BC_PP_0.omx. Fri Jul 14 11:01:25 PDT 2017 Loading spectra for 20170512_BC_PP_0.omx. Fri Jul 14 11:01:25 PDT 2017 Collecting peptides to map. Fri Jul 14 11:01:25 PDT 2017 Mapping peptides to proteins. Fri Jul 14 11:01:25 PDT 2017 Importing PSMs from 20170512_BC_PP_0.omx Fri Jul 14 11:01:28 PDT 2017 628 identified spectra (2.6%) did not present a valid peptide. Fri Jul 14 11:01:28 PDT 2017 17520 of the best scoring peptides were excluded by the import filters: Fri Jul 14 11:01:28 PDT 2017 - 84.0% peptide length less than 8 or greater than 30. Fri Jul 14 11:01:28 PDT 2017 - 16.0% peptide presenting high mass or isotopic deviation. Fri Jul 14 11:01:28 PDT 2017 Parsing 20170512_BC_PP_0.t.xml. Fri Jul 14 11:02:37 PDT 2017 Loading spectra for 20170512_BC_PP_0.t.xml. Fri Jul 14 11:02:37 PDT 2017 Collecting peptides to map. Fri Jul 14 11:02:38 PDT 2017 Mapping peptides to proteins. Fri Jul 14 11:02:39 PDT 2017 Importing PSMs from 20170512_BC_PP_0.t.xml Fri Jul 14 11:02:39 PDT 2017 9691 identified spectra (24.4%) did not present a valid peptide. Fri Jul 14 11:02:39 PDT 2017 14128 of the best scoring peptides were excluded by the import filters: Fri Jul 14 11:02:39 PDT 2017 - 31.3% peptide length less than 8 or greater than 30. Fri Jul 14 11:02:39 PDT 2017 - 64.1% peptide presenting high mass or isotopic deviation. Fri Jul 14 11:02:39 PDT 2017 - 4.5% unrecognized modifications. Fri Jul 14 11:02:39 PDT 2017 File import completed. 107115 first hits imported (538375 secondary) from 52232 spectra. Fri Jul 14 11:02:39 PDT 2017 [90002 first hits passed the initial filtering] Fri Jul 14 11:02:39 PDT 2017 Computing assumptions probabilities. Fri Jul 14 11:02:40 PDT 2017 Saving assumptions probabilities. Fri Jul 14 11:02:42 PDT 2017 Selecting best peptide per spectrum. Fri Jul 14 11:02:53 PDT 2017 Computing PSM probabilities. Fri Jul 14 11:02:53 PDT 2017 Scoring PTMs in PSMs (D-score and PhosphoRS) Fri Jul 14 11:05:41 PDT 2017 Resolving peptide inference issues. Fri Jul 14 11:06:18 PDT 2017 Saving probabilities, building peptides and proteins. Fri Jul 14 11:16:41 PDT 2017 Simplifying protein groups. Fri Jul 14 11:16:52 PDT 2017 31 unlikely protein mappings found: Fri Jul 14 11:16:52 PDT 2017 - 19 protein groups supported by non-enzymatic shared peptides. Fri Jul 14 11:16:52 PDT 2017 - 18 groups explained by a simpler group. Fri Jul 14 11:16:52 PDT 2017 Note: a group can present combinations of these criteria. Fri Jul 14 11:16:52 PDT 2017 Generating peptide map. Fri Jul 14 11:22:05 PDT 2017 Computing peptide probabilities. Fri Jul 14 11:22:05 PDT 2017 Saving peptide probabilities. Fri Jul 14 11:28:13 PDT 2017 Generating protein map. Fri Jul 14 11:30:19 PDT 2017 Resolving protein inference issues, inferring peptide and protein PI status. Fri Jul 14 12:06:23 PDT 2017 62 conflicts resolved. 14 protein groups remaining (0 suspicious). Fri Jul 14 12:06:23 PDT 2017 Correcting protein probabilities. Fri Jul 14 12:06:23 PDT 2017 Saving protein probabilities. Fri Jul 14 12:10:47 PDT 2017 Validating identifications at 1% FDR, quality control of matches. Fri Jul 14 12:23:25 PDT 2017 An error occurred: Object 20170512_BC_PP_0.mgf_cus_20170512_BC_PP_0.54911.54911.3 corresponding to entry PPvsDR_concise_PPvsDR_concise_0714_0_idccs"PeptideShaker|0_20170512_BC_PP_0.mgf_psm_parameters"_ccs_20170512_BC_PP_0.mgf_cus_20170512_BC_PP_0.54911.54911.3 not found in cache when saving. Fri Jul 14 12:23:25 PDT 2017 Please contact the developers ( https://github.com/compomics/peptide-shaker/issues).

Fri Jul 14 12:23:25 PDT 2017 Importing Data Canceled! Fri Jul 14 12:23:25 PDT 2017 An error occurred: null Fri Jul 14 12:23:25 PDT 2017 Please contact the developers ( https://github.com/compomics/peptide-shaker/issues).

On Fri, Jul 14, 2017 at 3:13 PM, Harald Barsnes notifications@github.com wrote:

Hi Joe,

I'm using the 1.16.11 version as well. Windows 10 and for this test I only gave PeptideShaker 4 GB of memory.

The snapshot of the error message is attached below.

The snapshot did not make it through the GitHub issue system I'm afraid. You have to upload them via the issue tracker, adding as attachment to the email does not work.

Best regards, Harald

— You are receiving this because you authored the thread. Reply to this email directly, view it on GitHub https://github.com/compomics/peptide-shaker/issues/274#issuecomment-315480474, or mute the thread https://github.com/notifications/unsubscribe-auth/AJ8rV_83ME6VQ3M9ZzvvzsP_q9HnTOejks5sN-fzgaJpZM4OWI3e .

hbarsnes commented 7 years ago

Hi Joe,

Sorry for the late reply. Seems like you are still having problems with the backend which I cannot reproduce on my side. Could you try loading the data for only one of the search engines alone to see if that works?

We are currently working on redesigning our backend which will hopefully also fix the problems you are having. This will however take a bit more time to complete.

Best regards, Harald

joeyuzh commented 7 years ago

Hi Harald,

I tried to just use only 'X!Tandem' this time. However, when loading the result in PeptideShaker, the following error message pops up.

Regards,

Joe

Tue Jul 18 20:56:05 PDT 2017 Validating identifications at 1% FDR, quality control of matches. Tue Jul 18 20:56:07 PDT 2017 Scoring PTMs in peptides.

Tue Jul 18 20:56:08 PDT 2017 Importing Data Canceled! Tue Jul 18 20:56:08 PDT 2017 An error occurred while loading the identification files: Tue Jul 18 20:56:08 PDT 2017 Attempting to create duplicate peptide key: MEPARAHWVK_-23.01598441731-ATAA-7_19.98982923912-ATAA-8_33.96909379454-ATAA-143.00581365643 from peptide MEPARAHWVK-23.01598441731-ATAA-7_15.99491461956-ATAA-1_19.98982923912-ATAA-8_33.96909379454-ATAA-1_43.00581365643.

hbarsnes commented 7 years ago

Hi Joe,

This new error is related to an issue with the PTM mapping that my colleague @mvaudel is working on. I've therefore forwarded your data to him for further testing. Hopefully your additional data set will make it easier for him to locate and fix the issue.

Best regards, Harald

joeyuzh commented 7 years ago

Hi Harald,

Thanks for your reply. Recently, I have been playing with search settings in SearchGUI. One thing pretty interesting that I noticed was that If I included 'Dioxidation of M' and 'Oxidation of M' in the variable modifications at the the same time. PeptideShaker would have trouble opening the file from SearchGUI with the error message:

"Mon Jul 24 14:56:26 PDT 2017 An error occurred while loading the identification files: Mon Jul 24 14:56:26 PDT 2017 Attempting to create duplicate peptide key: NKKAAEEAAK_43.00581365643-ATAA-2_43.00581365643-ATAA-3 from peptide NKKAAEEAAK_15.99491461956-ATAA-3_43.00581365643-ATAA-2_43.00581365643-ATAA-3."

This is also true for having 'Oxidation of K' and 'Carbamilation of K' in variable modification list at the same time. It seems to me that PeptideShaker doesn't allow different modifications for the same amino acid. Is that true?

Also, when I add the user defined modifications in the search settings, can I add something like 'oxidation of RYHW' or should I separate them into four independent items such as 'oxidation of R', 'oxidation of Y' and so on?

Regards,

Joe

mvaudel commented 7 years ago

Dear Joe,

You are correct that PeptideShaker does not "stack" modifications, but it is fine to provide multiple modifications on a single amino acid, the tool should be able to distribute them across the different sites. It is this site inference code that is causing problems at the moment. I am looking into it but it is not as simple as it seems, so will need a bit more time :) Also, we are currently refactoring our back-end code so I cannot make too big changes on the master branch at the moment.

As for the question regarding the different ways of designing an oxidation both should give the same results. I generally prefer having one modification per amino acid to make things cleaner :) Unless you have a substantial proportion of modified peptides, I would try to keep the modification list as short as possible to reduce the prevalence of false positives.

Hope this helps,

Marc

ulrich-eckhard commented 7 years ago

Hey Mark,

I am not sure if it fully fits, but let's see. A while back I quickly tried to look at a dimethyl experiment of a former colleague. i am aware PeptideShaker (PS) isn't for MS1 quantitation, but we just wanted to see if the labeling worked equally well and if I get roughly the same number of IDs for the two channels.

So I tried different set-ups, but I never managed to combine the data properly in PS. As a side note, I used comet-only in SearchGUI. So I tried e.g. two separate searches, and then combine them in PS providing a parameter file that had all modifications in it. But it failed. error message was I think e.g.: "Attempting to put two fixed modifications of different masses (34.xxx, 28.xxx) at position X in peptide Y" (so it didn't want to keep it as "separate" modification specific peptides, but instead tried to make it "one"). so i did two variable searches, so once dm0-var and one dm6-var (too avoid cross-modification),and used an dm0/dm6-variable parameter file for import. and PS managed to load the data, but only one set of dimethylation survived while both sets were in the parameter file provided for the import. but i defenitely don't remember all the details. i could try to redo something here if you have some good instructions what to try first ...

but as said, not sure if it really fits here. But per se it would be cool if something like that would work! :-) cheers u

On Tue, Jul 25, 2017 at 9:13 AM, Marc Vaudel notifications@github.com wrote:

Dear Joe,

You are correct that PeptideShaker does not "stack" modifications, but it is fine to provide multiple modifications on a single amino acid, the tool should be able to distribute them across the different sites. It is this site inference code that is causing problems at the moment. I am looking into it but it is not as simple as it seems, so will need a bit more time :) Also, we are currently refactoring our back-end code so I cannot make too big changes on the master branch at the moment.

As for the question regarding the different ways of designing an oxidation both should give the same results. I generally prefer having one modification per amino acid to make things cleaner :) Unless you have a substantial proportion of modified peptides, I would try to keep the modification list as short as possible to reduce the prevalence of false positives.

Hope this helps,

Marc

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mvaudel commented 7 years ago

Hi Ulrich,

Mmmmh that is also peculiar. PeptideShaker should be able to import peptides with both modifications variable. I will see if this is related.

Thanks for the input,

Marc

ulrich-eckhard commented 7 years ago

Hey Marc,

as said - is from a while back - a month or so, so don't remember all the details and didn't take proper notes. But still have the data and can do some testing later this week if you want me to try some things out ... e.g. using a different search engine or so. Or if i can reproduce the problem as there is for sure another PS version out there, and maybe it is a good idea to try again with a fresh mind.

just let me know, u

On Tue, Jul 25, 2017 at 11:15 AM, Marc Vaudel notifications@github.com wrote:

Hi Ulrich,

Mmmmh that is also peculiar. PeptideShaker should be able to import peptides with both modifications variable. I will see if this is related.

Thanks for the input,

Marc

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mvaudel commented 7 years ago

Sure. we will need help with testing when the new versions are out, I will send a mail on the mailing list when we are done refactoring our back-end :)

ulrich-eckhard commented 7 years ago

looking forward to it! :-)

On Tue, Jul 25, 2017 at 1:14 PM, Marc Vaudel notifications@github.com wrote:

Sure. we will need help with testing when the new versions are out, I will send a mail on the mailing list when we are done refactoring our back-end :)

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joeyuzh commented 7 years ago

Hi Mark,

Thanks for your reply. My project is interested in exploring the oxidized proteome under different experimental conditions. Accordingly, I try to input all possible oxidized modifications in Unimod (> 20) in the search settings. If some modifications cannot exist at the same, I will see if I can separate them into different search setting groups and sum up the search results at the end. But if PeptideShake can process them at one time, it will be perfect :)

Regards,

Joe

wagnerfontes2 commented 7 years ago

Hi, I think my issue is similar to the one that started this discussion, then I´m posting here. After running SearchGUI for a long while it tried to send the results to PeptideShaker but it crashed with a simple message that the process was cancelled. Please find below the PeptideShaker log. Please let me know if you need any other files, since they are rather large.

Sun Jul 30 21:10:47 BRT 2017: PeptideShaker version 1.16.12. Memory given to the Java virtual machine: 38177603584. Total amount of memory in the Java virtual machine: 128974848. Free memory: 102100488. Java version: 1.8.0_141. 1714 script command tokens (C) 2009 Jmol Development Jmol Version: 12.0.43 2011-05-03 14:21 java.vendor: Oracle Corporation java.version: 1.8.0_141 os.name: Windows 7 memory: 63.2/163.1 processors available: 32 useCommandThread: false Reindexing: uniprot-human-reviewed-trypsin-june-2017_concatenated_target_decoy.fasta. Reindexing: qExactive01819.mgf. (changes in the file detected)

Mon Jul 31 18:38:45 BRT 2017: PeptideShaker version 1.16.12. Memory given to the Java virtual machine: 38177603584. Total amount of memory in the Java virtual machine: 128974848. Free memory: 95823296. Java version: 1.8.0_141.

Tue Aug 01 14:52:18 BRT 2017: PeptideShaker version 1.16.12. Memory given to the Java virtual machine: 38177603584. Total amount of memory in the Java virtual machine: 163053568. Free memory: 105271832. Java version: 1.8.0_141. 1714 script command tokens (C) 2009 Jmol Development Jmol Version: 12.0.43 2011-05-03 14:21 java.vendor: Oracle Corporation java.version: 1.8.0_141 os.name: Windows 7 memory: 63.3/231.2 processors available: 32 useCommandThread: false Exception in thread "AWT-EventQueue-0" java.lang.NullPointerException at eu.isas.peptideshaker.gui.tabpanels.ValidationPanel.resetThreshold(ValidationPanel.java:3227) at eu.isas.peptideshaker.gui.tabpanels.ValidationPanel.resetAllThresholds(ValidationPanel.java:3214) at eu.isas.peptideshaker.gui.tabpanels.ValidationPanel.thresholdResetJButtonActionPerformed(ValidationPanel.java:2184) at eu.isas.peptideshaker.gui.tabpanels.ValidationPanel.access$6500(ValidationPanel.java:59) at eu.isas.peptideshaker.gui.tabpanels.ValidationPanel$49.actionPerformed(ValidationPanel.java:1140) at javax.swing.AbstractButton.fireActionPerformed(Unknown Source) at javax.swing.AbstractButton$Handler.actionPerformed(Unknown Source) at javax.swing.DefaultButtonModel.fireActionPerformed(Unknown Source) at javax.swing.DefaultButtonModel.setPressed(Unknown Source) at javax.swing.plaf.basic.BasicButtonListener.mouseReleased(Unknown Source) at java.awt.AWTEventMulticaster.mouseReleased(Unknown Source) at java.awt.AWTEventMulticaster.mouseReleased(Unknown Source) at java.awt.Component.processMouseEvent(Unknown Source) at javax.swing.JComponent.processMouseEvent(Unknown Source) at java.awt.Component.processEvent(Unknown Source) at java.awt.Container.processEvent(Unknown Source) at java.awt.Component.dispatchEventImpl(Unknown Source) at java.awt.Container.dispatchEventImpl(Unknown Source) at java.awt.Component.dispatchEvent(Unknown Source) at java.awt.LightweightDispatcher.retargetMouseEvent(Unknown Source) at java.awt.LightweightDispatcher.processMouseEvent(Unknown Source) at java.awt.LightweightDispatcher.dispatchEvent(Unknown Source) at java.awt.Container.dispatchEventImpl(Unknown Source) at java.awt.Window.dispatchEventImpl(Unknown Source) at java.awt.Component.dispatchEvent(Unknown Source) at java.awt.EventQueue.dispatchEventImpl(Unknown Source) at java.awt.EventQueue.access$500(Unknown Source) at java.awt.EventQueue$3.run(Unknown Source) at java.awt.EventQueue$3.run(Unknown Source) at java.security.AccessController.doPrivileged(Native Method) at java.security.ProtectionDomain$JavaSecurityAccessImpl.doIntersectionPrivilege(Unknown Source) at java.security.ProtectionDomain$JavaSecurityAccessImpl.doIntersectionPrivilege(Unknown Source) at java.awt.EventQueue$4.run(Unknown Source) at java.awt.EventQueue$4.run(Unknown Source) at java.security.AccessController.doPrivileged(Native Method) at java.security.ProtectionDomain$JavaSecurityAccessImpl.doIntersectionPrivilege(Unknown Source) at java.awt.EventQueue.dispatchEvent(Unknown Source) at java.awt.EventDispatchThread.pumpOneEventForFilters(Unknown Source) at java.awt.EventDispatchThread.pumpEventsForFilter(Unknown Source) at java.awt.EventDispatchThread.pumpEventsForHierarchy(Unknown Source) at java.awt.EventDispatchThread.pumpEvents(Unknown Source) at java.awt.EventDispatchThread.pumpEvents(Unknown Source) at java.awt.EventDispatchThread.run(Unknown Source) Reindexing: Trinity.fasta.transdecoder.pep_adap_withdecoy_GUI.fasta. 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Wed Aug 23 23:35:43 BRT 2017: PeptideShaker version 1.16.12. Memory given to the Java virtual machine: 38177603584. Total amount of memory in the Java virtual machine: 128974848. Free memory: 113013544. Java version: 1.8.0_141. java.lang.IllegalArgumentException: Query error: Query ERROR: caught BioMart::Exception::Usage: Attribute uniprotswissprot NOT FOUND at com.compomics.util.experiment.biology.genes.GeneFactory.queryEnsembl(GeneFactory.java:484) at com.compomics.util.experiment.biology.genes.GeneFactory.downloadGoMappings(GeneFactory.java:379) at com.compomics.util.experiment.biology.genes.GeneFactory.downloadMappings(GeneFactory.java:912) at com.compomics.util.experiment.biology.genes.GeneFactory.getGeneMaps(GeneFactory.java:168) at eu.isas.peptideshaker.fileimport.FileImporter.importGenes(FileImporter.java:276) at eu.isas.peptideshaker.fileimport.FileImporter$IdProcessorFromFile.importFiles(FileImporter.java:453) at eu.isas.peptideshaker.fileimport.FileImporter.importFiles(FileImporter.java:151) at eu.isas.peptideshaker.PeptideShaker.importFiles(PeptideShaker.java:229) at eu.isas.peptideshaker.cmd.PeptideShakerCLI.createProject(PeptideShakerCLI.java:735) at eu.isas.peptideshaker.cmd.PeptideShakerCLI.call(PeptideShakerCLI.java:218) at eu.isas.peptideshaker.cmd.PeptideShakerCLI.main(PeptideShakerCLI.java:935) java.lang.NullPointerException at com.compomics.util.experiment.identification.protein_inference.fm_index.FMIndex.getProteinMappingWithoutVariants(FMIndex.java:4155) at com.compomics.util.experiment.identification.protein_inference.fm_index.FMIndex.getProteinMapping(FMIndex.java:3827) at com.compomics.util.experiment.identification.protein_inference.fm_index.FMIndex.getProteinMapping(FMIndex.java:3801) at eu.isas.peptideshaker.protein_inference.TagMapper.mapTagsForSpectrumMatch(TagMapper.java:192) at eu.isas.peptideshaker.protein_inference.TagMapper.access$000(TagMapper.java:54) at eu.isas.peptideshaker.protein_inference.TagMapper$SpectrumMatchTagMapperRunnable.run(TagMapper.java:380) at java.util.concurrent.Executors$RunnableAdapter.call(Unknown Source) at java.util.concurrent.FutureTask.run(Unknown Source) at java.util.concurrent.ThreadPoolExecutor.runWorker(Unknown Source) at java.util.concurrent.ThreadPoolExecutor$Worker.run(Unknown Source) at java.lang.Thread.run(Unknown Source)

Fri Aug 25 07:07:56 BRT 2017: PeptideShaker version 1.16.12. Memory given to the Java virtual machine: 38177603584. Total amount of memory in the Java virtual machine: 163053568. Free memory: 82948808. Java version: 1.8.0_141. 1714 script command tokens (C) 2009 Jmol Development Jmol Version: 12.0.43 2011-05-03 14:21 java.vendor: Oracle Corporation java.version: 1.8.0_141 os.name: Windows 7 memory: 74.6/231.2 processors available: 32 useCommandThread: false

Thanks, Wagner.

hbarsnes commented 7 years ago

Hi Wagner,

I don't really see how the issue you are having is related to the one in this thread? Could you please open a new issue where you explain a bit more about the problem and what kind if data you are trying to load? If you could also share with us the data you are trying to load in PeptideShaker that would be great.

Best regards, Harald

hbarsnes commented 4 years ago

This issue should hopefully be solved in the new beta releases of SearchGUI and PeptideShaker. Please see https://groups.google.com/forum/#!topic/peptide-shaker/1ecY0IyMOBM for more details.

If the issue still exists in the beta releases, please open a new issue at https://github.com/compomics/peptide-shaker-2.0-issue-tracker/issues.

If the issue has been solved, it would also be great of you could let us know by replying to this original issue and letting us know. :)

hbarsnes commented 4 years ago

This issue should hopefully be fixed in the new release that is available now. If not, please let us know by opening a new issue.