Failing on QC analysis with database #76
Comments
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Hi, Can you give me some more details? Are you using the command line version of RawTools? If so, what is the command you are feeding it? If you are using the GUI, can you tell me the options you are providing it? Can you let me know what the database is you are using? I assume you are using the latest version of RawTools? Do I understand correctly that this error only occurs if you specify a sequence database? If you don't try to do a search everything works as expected? Any info you can give that can help me reproduce the error in order to fix it is much appreciated. Chris |
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Hi, Im using the command line version of RawTools. the command I've used is:
I'm using the Uniprot UP000005640 human database Yes, I am using the latest version. And yes again, the issue only occurs when I sequence a database. I have just tried doing it with the Gui and turned up an error that reads like this: the setting I used are: I hope this help clarify my situation. Ilham |
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Hi, Thank you for this additional information. It appears the GUI and command line errors are different. I think we should just focus on the command line one for now. I am having trouble reproducing the error on my own system (for reference, I am running on a Windows machine with RawTools 2.0.7). If I use the same command you have on the command line using just a subset of the raw files you are using, my command executes fine: C:\Users\chughes\Documents\bccrc\softwareRepository\RawTools\rawtools207>RawTools.exe -d C:\Users\chughes\Documents\bccrc\softwareRepository\RawTools\raw-test-file\testingQc\ -qc C:\Users\chughes\Documents\bccrc\softwareRepository\RawTools\raw-test-file\testingQc\ -db C:\Users\chughes\Documents\bccrc\softwareRepository\RawTools\raw-test-file\testingQc\UP000005640_9606.fasta -fmods 57.0214@C -vmods 15.9949@M -X C:\Users\chughes\Documents\bccrc\softwareRepository\RawTools\tandem-win-17-02-01-4\tandem-win-17-02-01-4\bin
2 file(s) to process
Processing: C:\Users\chughes\Documents\bccrc\softwareRepository\RawTools\raw-test-file\testingQc\ch_23Aug2018_HeLa_Std_1.raw
Determing MS analysis order... Done!
Extracting scan indices: 100%
Checking for orphaned scans: 100%
================ Scan indexing report ================
Total scans in file: 11378
Scans linked: 11378
Orphan scans:
None!
All scans accounted for!
======================================================
Extracting reaction events: 100%
Extracting scan data: 100%
Extracting trailer extras: 100%
Extracting precursor masses: 100%
Extracting retention times: 100%
Analyzing precursor peaks: 100%
Calculating meta data
MS1 isolation interference
MS2 scan cycle density
Ion injection time
FAIMS voltages
Duty cycle
Intensity distribution
Summed intensities
Calculating metrics
Writing MGF file: 100%
Reading FASTA file
Writing target-decoy database: 100%
X! TANDEM Alanine (2017.2.1.4)
Loading spectra| (mgf).. loaded.
Spectra matching criteria = 2969
Starting threads .|.|.|.|.|.|.|.|.|.|.|.|.|.|. started.
Computing models:
testing 1 2 3 testing 1 2 3 testing 1 2 3
waiting for 23|45|6|7|8|9|10|11|12|13|14|15| done.
sequences modelled = 41 ks
Model refinement:
waiting for 2|3|4|5|6|7|8|9|10|11|12|13|14|15| done.
Merging results:
from 23456789101112131415
Creating report:
initial calculations ..... done.
sorting ..... done.
finding repeats ..... done.
evaluating results ..... done.
calculating expectations ..... done.
writing results ..... done.
Valid models = 2038
Sufficient PSMs and decoy hits detected for ID filtering.
Total number of unfiltered PSMs: 2960
Total number of decoy hits: 300
Total hits: 2960
Top decoy score: 30.8
Non-decoy hits: 2660
Non-decoy hits above top decoy score: 1906
Digestion efficiency: 0.707764952780693
Missed cleavage rate (/PSM): 0.322665267576076
IDrate: 0.635333333333333
15.9949@M modification frequency: 0.0107858243451464
Finished writing QC data to csv
QC data written to csv file.
QC file saved successfully
Elapsed time: 15.03 s
Processing: C:\Users\chughes\Documents\bccrc\softwareRepository\RawTools\raw-test-file\testingQc\ch_23Aug2018_HeLa_Std_2.raw
Determing MS analysis order... Done!
Extracting scan indices: 100%
Checking for orphaned scans: 100%
================ Scan indexing report ================
Total scans in file: 11525
Scans linked: 11525
Orphan scans:
None!
All scans accounted for!
======================================================
Extracting reaction events: 100%
Extracting scan data: 100%
Extracting trailer extras: 100%
Extracting precursor masses: 100%
Extracting retention times: 100%
Analyzing precursor peaks: 100%
Calculating meta data
MS1 isolation interference
MS2 scan cycle density
Ion injection time
FAIMS voltages
Duty cycle
Intensity distribution
Summed intensities
Calculating metrics
Writing MGF file: 100%
X! TANDEM Alanine (2017.2.1.4)
Loading spectra| (mgf).. loaded.
Spectra matching criteria = 2967
Starting threads .|.|.|.|.|.|.|.|.|.|.|.|.|.|. started.
Computing models:
testing 1 2 3 testing 1 2 3 testing 1 2 3
waiting for 2|3|45|6|7|8|9|10|11|12|13|14|15| done.
sequences modelled = 41 ks
Model refinement:
waiting for 2|3|4|5|6|7|8|9|10|11|12|13|14|15| done.
Merging results:
from 23456789101112131415
Creating report:
initial calculations ..... done.
sorting ..... done.
finding repeats ..... done.
evaluating results ..... done.
calculating expectations ..... done.
writing results ..... done.
Valid models = 2080
Sufficient PSMs and decoy hits detected for ID filtering.
Total number of unfiltered PSMs: 2959
Total number of decoy hits: 293
Total hits: 2959
Top decoy score: 30.05
Non-decoy hits: 2666
Non-decoy hits above top decoy score: 1997
Digestion efficiency: 0.72158237356034
Missed cleavage rate (/PSM): 0.302954431647471
IDrate: 0.665666666666667
15.9949@M modification frequency: 0.0258992805755396
Finished writing QC data to csv
QC data written to csv file.
QC file saved successfully
Elapsed time: 13.78 s
Time to process all 2 files: 00:00:28.8167614
C:\Users\chughes\Documents\bccrc\softwareRepository\RawTools\rawtools207>If I execute the same command in the GUI, it also works fine, so I am a bit at a loss here. When you run it from the command line, at what point during the execution does it fail? I can you provide me with the entire command line output (including your input command and any information RawTools spits out at all, perhaps running with and without a search). |
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Hi, As a note, I am also running windows, am using RawTools 2.0.7, and also actually only using a subset of the raw files I mentioned. with this, the results are created in the directory I specified When I run RawTools with the search, my output looks like this: Since the issue mentions not finding a file, I tried reinstalling it thinking that maybe i had done it wrong the first time. This didn't bear any fruit and i returned the exact same errors. Thanks again Ilham |
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If you check the QC directory, does it actually write the target decoy database? |
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It produces a new database in the same directory as the original
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It is a bit odd that your 'RawTools_custom_config.xml' file is in that directory and not in your xtandem/bin directory (or is this one just a copy?). Can you show me what your xtandem bin directory looks like? I assume the mgf is also written? Would you be willing to try an older version, just for a sanity check (version 2.0.4 is a good one to try). |
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I haven't made any copies of the "RawTools_custom_config.xml" file, so it should be the original. here's an image of the xtandem bin:
The only mgf file i could find was in the results folder. It only wrote for one of my input files: I'll give 2.0.4 a shot and let you know how that goes as well. |
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Just did it with version 2.0.4 and it worked! I've have to run a few more sets of data to be sure, but the test files i was using have been successfully processed this is what the console spit out: I don't want to get my hopes up too early though. I'll reach out again if I need anymore help |
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Hmm. I will have to try and track down what was happening in 2.0.7 at some point. It is odd though as nothing really changed in the QC pipeline between 2.0.4 and 2.0.7. I will have to think about it. |
Hi, when i run the basic QC analysis, the program produced a output as intended. But when I try to do the QC with the database, the program fails and returns this error
Unhandled Exception: System.ComponentModel.Win32Exception: The system cannot find the file specified at System.Diagnostics.Process.StartWithCreateProcess(ProcessStartInfo startInfo) at RawTools.Utilities.ConsoleUtils.VoidBash(String cmd, String args) in C:\Users\chughes\Documents\bccrc\softwareRepository\RawTools\rawtoolsDev\RawTools\Utilities\Utilities.cs:line 109 at RawTools.QC.Search.RunSearch(WorkflowParameters parameters, MethodDataContainer methodData, String rawFileName) in C:\Users\chughes\Documents\bccrc\softwareRepository\RawTools\rawtoolsDev\RawTools\QC\RunSearch.cs:line 63 at RawTools.WorkFlows.WorkFlowsDDA.UniversalDDA(IRawFileThreadManager rawFileThreadManager, WorkflowParameters parameters, QcDataCollection qcDataCollection) in C:\Users\chughes\Documents\bccrc\softwareRepository\RawTools\rawtoolsDev\RawTools\WorkFlows\DDA.cs:line 188 at RawTools.Program.Run(Dictionary2 opts) in C:\Users\chughes\Documents\bccrc\softwareRepository\RawTools\rawtoolsDev\RawTools\Program.cs:line 243at RawTools.Program.Main(String[] args) in C:\Users\chughes\Documents\bccrc\softwareRepository\RawTools\rawtoolsDev\RawTools\Program.cs:line 73`
I'm new to software like this and was hoping for some help understanding the issue.
*note i am using raw files from the EBI Pride database project PXD011070 (the files suggested in the guide you have)
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