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Nesvilab / IonQuant

A label free quantification tool.
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IonQuant filtering: Protein not found in psm.tsv #12

Closed grosenberger closed 3 years ago

grosenberger commented 3 years ago

Dear IonQuant developers,

I'm trying to set up a workflow based on MSFragger, Philosopher (PeptideProphet, iProphet & ProteinProphet) & IonQuant. Essentially N MSFragger searches are conducted, followed by N PeptideProphet, 1 iProphet, 1 ProteinProphet & 1 philosopher filter runs. Then I use the output N PeptideProphet & 1 psm.tsv files together with the original mzXML files to run IonQuant. Most steps work out, however at the last step, I encounter a problem:

2020-11-09 09:19:25 [ERROR] - Protein sp|Q15415|RBY1F_HUMAN from data_quant/protein.tsv cannot be found from any psm.tsv.

I use the following command to run IonQuant:

java -Xmx14G -jar ./IonQuant-1.4.4.jar --threads 4 --psm data_quant/psm.tsv --mbr 0 --ionfdr 0.01 --peptidefdr 0.01 --proteinfdr 0.01 --multidir data_quant --specdir data_search data_search/*.pepXML

Is there anything I need to do to make it work? Thanks for your help!

Best regards, George

fcyu commented 3 years ago

Hi George,

Thanks for your feedback. This is a known issue. I released 1.4.6 that should fix it. Could you please try and see if it works for you?

Best,

Fengchao

grosenberger commented 3 years ago

Hi Fengchao,

Excellent, thank you for the fast patch! This seems to have solved the issue and IonQuant, including MBR runs through. However, I have found that some runs are excluded in the final report. Is there a parameter that needs to be tweaked to include those as well?

Best regards, George

fcyu commented 3 years ago

Hi George,

I don't think we have any parameter excluding runs. Could you please elaborate more and send us the log if needed?

Best,

Fengchao

grosenberger commented 3 years ago

Hi Fengchao,

Modifying the parameters didn't help. The following message might provide some clues:

2020-11-12 21:32:35 [WARNING] - There is no mbr file for run 043_Prot109_D19. May be because it failed in MBR.

Indeed this and other runs don't have any generated _quant.csv or _model.png files associated with them and the following error might indicate that the search was not successful, right?

2020-11-12 21:07:33 [INFO] - Matching-between-runs: 106_Prot109_D19...
2020-11-12 21:07:50 [INFO] - Calculating correlations between all other runs...
2020-11-12 21:12:05 [INFO] - Correlation with 099_Prot109_D19: 0.0013531799
2020-11-12 21:12:05 [INFO] - Correlation with 109_Prot109_D19: 8.069397E-4
2020-11-12 21:12:05 [INFO] - Correlation with 105_Prot109_D19: 6.5220933E-4
2020-11-12 21:12:05 [INFO] - Correlation with 098_Prot109_D19: 5.8823527E-4
2020-11-12 21:12:05 [INFO] - Correlation with 085_Prot109_D19: 4.229522E-4
2020-11-12 21:12:05 [INFO] - Correlation with 071_Prot109_D19: 4.014452E-4
2020-11-12 21:12:05 [INFO] - Correlation with 033_Prot109_D19: 3.6127167E-4
2020-11-12 21:12:05 [INFO] - Correlation with 084_Prot109_D19: 2.889895E-4
2020-11-12 21:12:05 [INFO] - Correlation with 004_Prot109f_D19: 2.4169184E-4
2020-11-12 21:12:05 [INFO] - Correlation with 077_Prot109_D19: 2.4163345E-4
2020-11-12 21:12:05 [INFO] - Correlation with 075_Prot109_D19: 2.392917E-4
2020-11-12 21:12:05 [INFO] - Correlation with 013_Prot109f_D19: 2.3542953E-4
2020-11-12 21:12:05 [INFO] - Correlation with 074_Prot109_D19: 2.1850759E-4
2020-11-12 21:12:05 [INFO] - Correlation with 083_Prot109_D19: 2.184254E-4
2020-11-12 21:12:05 [INFO] - Correlation with 090_Prot109_D19: 2.034381E-4
2020-11-12 21:12:05 [INFO] - Correlation with 020_Prot109f_D19: 1.9406171E-4
2020-11-12 21:12:05 [INFO] - Correlation with 080_Prot109_D19: 1.8271514E-4
2020-11-12 21:12:05 [INFO] - Correlation with 007_Prot109f_D19: 1.8125793E-4
2020-11-12 21:12:05 [INFO] - Correlation with 026_Prot109_D19: 1.631987E-4
2020-11-12 21:12:05 [INFO] - Correlation with 017_Prot109ff_D19: 1.6095284E-4
2020-11-12 21:12:05 [INFO] - Correlation with 027_Prot109_D19: 1.5555728E-4
2020-11-12 21:12:05 [INFO] - Correlation with 022_Prot109_D19: 1.4716704E-4
2020-11-12 21:12:05 [INFO] - Correlation with 018_Prot109f_D19: 1.1517199E-4
2020-11-12 21:12:05 [INFO] - Correlation with 021_Prot109f_D19: 7.431629E-5
2020-11-12 21:12:05 [INFO] - Correlation with 019_Prot109ff_D19: 6.743392E-5
2020-11-12 21:12:05 [INFO] - Correlation with 062_Prot109_D19: 0.0
2020-11-12 21:12:05 [INFO] - Correlation with 038_Prot109_D19: 0.0
2020-11-12 21:12:05 [INFO] - Correlation with 031_Prot109_D19: 0.0
2020-11-12 21:12:05 [INFO] - Correlation with 030_Prot109_D19: 0.0
2020-11-12 21:12:05 [INFO] - Correlation with 024_Prot109_D19: 0.0
2020-11-12 21:12:05 [INFO] - Correlation with 023_Prot109_D19: 0.0
2020-11-12 21:12:05 [INFO] - Correlation with 108_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 107_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 097_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 096_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 095_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 094_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 093_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 092_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 091_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 089_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 088_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 087_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 086_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 082_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 081_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 079_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 078_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 076_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 073_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 072_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 064_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 063_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 061_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 060_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 058_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 050_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 049_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 048_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 047_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 046_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 045_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 042_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 041_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 040_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 039_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 037_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 036_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 035_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 034_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Correlation with 032_Prot109_D19: NaN
2020-11-12 21:12:05 [INFO] - Reading index from  106_Prot109_D19.quantindex
2020-11-12 21:12:06 [INFO] - Mapping ions from 099_Prot109_D19 to 106_Prot109_D19...
2020-11-12 21:12:23 [INFO] - Mapping ions from 109_Prot109_D19 to 106_Prot109_D19...
2020-11-12 21:12:41 [INFO] - Mapping ions from 105_Prot109_D19 to 106_Prot109_D19...
2020-11-12 21:12:59 [INFO] - Mapping ions from 098_Prot109_D19 to 106_Prot109_D19...
2020-11-12 21:13:18 [INFO] - Mapping ions from 085_Prot109_D19 to 106_Prot109_D19...
2020-11-12 21:13:36 [WARNING] - Run 106_Prot109_D19 got nothing matched.
fcyu commented 3 years ago

Hi George,

It looks like there is not much overlapped ions between 043_Prot109_D19 and other runs, so the MBR failed. Please note that there need to be a certain amount of overlapping to perform the RT and/or ion mobility alignment for MBR.

Best,

Fengchao

grosenberger commented 3 years ago

Hi Fengchao,

Thanks for the advice. I will try to tweak the database search settings. Probably there are some issues that need to be fixed there regarding the parameter selection. IonQuant looks like a a fantastic tool and I'm excited to see how it performs on the dataset that I'm analysing now.

Best regards, George