Why Is MSFragger Called Instead of MSFragger-DIA When Running DIA_SpecLib_Quant via FragPipe CLI? #2656
Description
I am using FragPipe in command-line mode within a Nextflow pipeline to perform identification on DIA data. In the FragPipe GUI, I selected the DIA_SpecLib_Quant workflow and changed “Filetype to Convert” from pepXML to psm.tsv, then exported this workflow.
In Nextflow, I first converted the DIA .raw files to .mzML using msconvert, and then ran FragPipe in command-line mode with the exported workflow and the .mzML files. The pipeline completed successfully; however, I noticed that the output files look very similar to those from a DDA run. In particular, it appears that MSFragger was called instead of MSFragger-DIA, which does not match the expected behavior of the DIA_SpecLib_Quant workflow in the FragPipe GUI.
I am not sure where the issue originates and would appreciate any guidance on what might be missing or misconfigured in this setup.
There is some of the information in fragpipe.stdout.log
System OS: Linux, Architecture: amd64
Java Info: 17.0.15, OpenJDK 64-Bit Server VM, Ubuntu
.NET Core Info: 6.0.36
Version info:
FragPipe version 23.0
DIA-Umpire version 2.3.2
diaTracer version 1.3.3
MSFragger version 4.3
Crystal-C version 1.5.8
MSBooster version 1.3.9
Percolator version 3.7.1
Philosopher version 5.1.1
PTM-Shepherd version 3.0.1
IonQuant version 1.11.11
TMT-Integrator version 6.1.1
EasyPQP version 0.1.52
DIA-NN version 1.8.2 Beta 8
Skyline version N/A
Pandas version 2.2.3
Numpy version 1.26.4
LCMS files:
Experiment/Group:
(if "spectral library generation" is enabled, all files will be analyzed together)
- ./20240228_2024-17_Maarten_04_DIA_T1.mzML DIA
26 commands to execute:
CheckCentroid
java -Xmx37G -cp /fragpipe_bin/fragpipe-23.0/fragpipe-23.0/lib/fragpipe-23.0.jar:/fragpipe_bin/fragpipe-23.0/fragpipe-23.0/tools/batmass-io-1.35.1.jar org.nesvilab.fragpipe.util.CheckCentroid /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/20240228_2024-17_Maarten_04_DIA_T1.mzML 35
WorkspaceCleanInit [Work dir: /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940]
/fragpipe_bin/fragpipe-23.0/fragpipe-23.0/tools/Philosopher/philosopher-v5.1.1 workspace --clean --nocheck
WorkspaceCleanInit [Work dir: /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940]
/fragpipe_bin/fragpipe-23.0/fragpipe-23.0/tools/Philosopher/philosopher-v5.1.1 workspace --init --nocheck --temp /tmp/252d2d9b-db43-46f2-a8eb-7647897e10bf
MSFragger [Work dir: /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940]
java -jar -Dfile.encoding=UTF-8 -Xmx37G /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/tools_dir/MSFragger/MSFragger-4.3/MSFragger-4.3.jar /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/fragger_dia.params ./20240228_2024-17_Maarten_04_DIA_T1.mzML
MSFragger move pepxml
java -cp /fragpipe_bin/fragpipe-23.0/fragpipe-23.0/lib/fragpipe-23.0.jar:/fragpipe_bin/fragpipe-23.0/fragpipe-23.0/lib/batmass-io-1.35.1.jar org.nesvilab.utils.FileMove --no-err /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/20240228_2024-17_Maarten_04_DIA_T1_rank1.pepXML /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/20240228_2024-17_Maarten_04_DIA_T1_rank1.pepXML
MSFragger move pepxml
java -cp /fragpipe_bin/fragpipe-23.0/fragpipe-23.0/lib/fragpipe-23.0.jar:/fragpipe_bin/fragpipe-23.0/fragpipe-23.0/lib/batmass-io-1.35.1.jar org.nesvilab.utils.FileMove --no-err /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/20240228_2024-17_Maarten_04_DIA_T1_rank2.pepXML /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/20240228_2024-17_Maarten_04_DIA_T1_rank2.pepXML
MSFragger move pepxml
java -cp /fragpipe_bin/fragpipe-23.0/fragpipe-23.0/lib/fragpipe-23.0.jar:/fragpipe_bin/fragpipe-23.0/fragpipe-23.0/lib/batmass-io-1.35.1.jar org.nesvilab.utils.FileMove --no-err /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/20240228_2024-17_Maarten_04_DIA_T1_rank3.pepXML /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/20240228_2024-17_Maarten_04_DIA_T1_rank3.pepXML
MSFragger move pepxml
java -cp /fragpipe_bin/fragpipe-23.0/fragpipe-23.0/lib/fragpipe-23.0.jar:/fragpipe_bin/fragpipe-23.0/fragpipe-23.0/lib/batmass-io-1.35.1.jar org.nesvilab.utils.FileMove --no-err /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/20240228_2024-17_Maarten_04_DIA_T1_rank4.pepXML /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/20240228_2024-17_Maarten_04_DIA_T1_rank4.pepXML
MSFragger move pepxml
java -cp /fragpipe_bin/fragpipe-23.0/fragpipe-23.0/lib/fragpipe-23.0.jar:/fragpipe_bin/fragpipe-23.0/fragpipe-23.0/lib/batmass-io-1.35.1.jar org.nesvilab.utils.FileMove --no-err /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/20240228_2024-17_Maarten_04_DIA_T1_rank5.pepXML /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/20240228_2024-17_Maarten_04_DIA_T1_rank5.pepXML
MSFragger move pin
java -cp /fragpipe_bin/fragpipe-23.0/fragpipe-23.0/lib/fragpipe-23.0.jar:/fragpipe_bin/fragpipe-23.0/fragpipe-23.0/lib/batmass-io-1.35.1.jar org.nesvilab.utils.FileMove --no-err /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/20240228_2024-17_Maarten_04_DIA_T1.pin /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/20240228_2024-17_Maarten_04_DIA_T1.pin
MSBooster [Work dir: /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940]
java -Djava.awt.headless=true -Xmx37G -cp /fragpipe_bin/fragpipe-23.0/fragpipe-23.0/tools/MSBooster-1.3.9.jar:/fragpipe_bin/fragpipe-23.0/fragpipe-23.0/tools/batmass-io-1.35.1.jar mainsteps.MainClass --paramsList /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/msbooster_params.txt
Percolator [Work dir: /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940]
/fragpipe_bin/fragpipe-23.0/fragpipe-23.0/tools/percolator_3_7_1/linux/percolator --no-terminate --post-processing-tdc --subset-max-train 500000 --num-threads 35 --results-peptides 20240228_2024-17_Maarten_04_DIA_T1_percolator_target_psms.tsv --decoy-results-peptides 20240228_2024-17_Maarten_04_DIA_T1_percolator_decoy_psms.tsv --protein-decoy-pattern rev_ 20240228_2024-17_Maarten_04_DIA_T1_edited.pin
Percolator: Convert to pepxml [Work dir: /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940]
java -cp /fragpipe_bin/fragpipe-23.0/fragpipe-23.0/lib/* org.nesvilab.fragpipe.tools.percolator.PercolatorOutputToPepXML 20240228_2024-17_Maarten_04_DIA_T1.pin 20240228_2024-17_Maarten_04_DIA_T1 20240228_2024-17_Maarten_04_DIA_T1_percolator_target_psms.tsv 20240228_2024-17_Maarten_04_DIA_T1_percolator_decoy_psms.tsv interact-20240228_2024-17_Maarten_04_DIA_T1 DIA 0.7 /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/20240228_2024-17_Maarten_04_DIA_T1.mzML
Percolator delete temp
java -cp /fragpipe_bin/fragpipe-23.0/fragpipe-23.0/lib/fragpipe-23.0.jar org.nesvilab.utils.FileDelete /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/20240228_2024-17_Maarten_04_DIA_T1_percolator_target_psms.tsv
Percolator delete temp
java -cp /fragpipe_bin/fragpipe-23.0/fragpipe-23.0/lib/fragpipe-23.0.jar org.nesvilab.utils.FileDelete /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/20240228_2024-17_Maarten_04_DIA_T1_percolator_decoy_psms.tsv
ProteinProphet [Work dir: /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940]
/fragpipe_bin/fragpipe-23.0/fragpipe-23.0/tools/Philosopher/philosopher-v5.1.1 proteinprophet --maxppmdiff 2000000 --minprob 0.5 --output combined /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/filelist_proteinprophet.txt
PhilosopherDbAnnotate [Work dir: /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940]
/fragpipe_bin/fragpipe-23.0/fragpipe-23.0/tools/Philosopher/philosopher-v5.1.1 database --annotate /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/2026-02-04-decoys-contam-2025-11-27__14-31-59.f70db2649f5e8c3c1331271851956fd7.merged.fas.fas --prefix rev_
PhilosopherFilter [Work dir: /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940]
/fragpipe_bin/fragpipe-23.0/fragpipe-23.0/tools/Philosopher/philosopher-v5.1.1 filter --picked --prot 0.01 --minPepLen 8 --tag rev_ --pepxml /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940 --protxml /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/combined.prot.xml --razor
PhilosopherReport [Work dir: /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940]
/fragpipe_bin/fragpipe-23.0/fragpipe-23.0/tools/Philosopher/philosopher-v5.1.1 report
WorkspaceClean [Work dir: /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940]
/fragpipe_bin/fragpipe-23.0/fragpipe-23.0/tools/Philosopher/philosopher-v5.1.1 workspace --clean --nocheck
SpecLibGen [Work dir: /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940]
python3 -u /fragpipe_bin/fragpipe-23.0/fragpipe-23.0/tools/speclib/gen_con_spec_lib.py /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/2026-02-04-decoys-contam-2025-11-27__14-31-59.f70db2649f5e8c3c1331271851956fd7.merged.fas.fas /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940 unused /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940 True unused use_easypqp noiRT:noIM 35 "--max_delta_unimod 0.02 --max_delta_ppm 15.0 --fragment_types ['b','y',] --decoy_prefix rev_ --max_glycan_q 1.0" "--rt_lowess_fraction 0.0" delete_intermediate_files /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/filelist_speclibgen.txt
DIA-Quant run DIA-NN [Work dir: /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940]
/fragpipe_bin/fragpipe-23.0/fragpipe-23.0/tools/diann/1.8.2_beta_8/linux/diann-1.8.1.8 --lib library.tsv --threads 35 --verbose 1 --out dia-quant-output/report.tsv --qvalue 0.01 --matrix-qvalue 0.01 --matrices --no-prot-inf --smart-profiling --no-quant-files --peak-center --no-ifs-removal --report-lib-info --cfg /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/filelist_diann.txt--
DIA-Quant move and rename speclib for skyline
java -cp /fragpipe_bin/fragpipe-23.0/fragpipe-23.0/lib/fragpipe-23.0.jar org.nesvilab.utils.FileMoveWithExtension /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940 .speclib /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/dia-quant-output/report-tsv.speclib
DIA-Quant propagate information [Work dir: /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/dia-quant-output]
java -cp /fragpipe_bin/fragpipe-23.0/fragpipe-23.0/tools/batmass-io-1.35.1.jar:/fragpipe_bin/fragpipe-23.0/fragpipe-23.0/lib/fragpipe-23.0.jar org.nesvilab.fragpipe.tools.diann.Propagation /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940 - - -
DIA-Quant convert DIA-NN output to MSstats.csv [Work dir: /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/dia-quant-output]
java -cp /fragpipe_bin/fragpipe-23.0/fragpipe-23.0/lib/commons-io-2.18.0.jar:/fragpipe_bin/fragpipe-23.0/fragpipe-23.0/lib/fragpipe-23.0.jar org.nesvilab.fragpipe.tools.diann.DiannToMsstats report.tsv ./ /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/psm.tsv 0.01 1 0.01 0.01 /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/fragpipe-files.fp-manifest
DIA-Quant generate site reports [Work dir: /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/dia-quant-output]
java -jar /fragpipe_bin/fragpipe-23.0/fragpipe-23.0/tools/LFQ-SiteReporter-1.0.3.jar -pr /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/dia-quant-output/report.tsv -psm /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/psm.tsv -out_dir /home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/dia-quant-output -mod_tag -min_site_prob 0.75
Execution order:
Cmd: [START], Work dir: [/home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940]
Cmd: [CheckCentroid], Work dir: [/home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940]
Cmd: [WorkspaceCleanInit], Work dir: [/home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940]
Cmd: [MSFragger], Work dir: [/home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940]
Cmd: [MSBooster], Work dir: [/home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940]
Cmd: [Percolator], Work dir: [/home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940]
Cmd: [ProteinProphet], Work dir: [/home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940]
Cmd: [PhilosopherDbAnnotate], Work dir: [/home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940]
Cmd: [PhilosopherFilter], Work dir: [/home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940]
Cmd: [PhilosopherReport], Work dir: [/home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940]
Cmd: [WorkspaceClean], Work dir: [/home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940]
Cmd: [SpecLibGen], Work dir: [/home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940]
Cmd: [DIA-Quant], Work dir: [/home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940]
>0000349febc09c9ceee824616a5a1a28
>36dde281a6c0ff1301ea56209a35eac4
>6de292cb70c9a38b804570dc7c658e6f
>ENSP00000257567.2 pep chromosome:GRCh38:12:114670255:114684175:-1 gene:ENSG00000135111.16 transcript:ENST00000349155.7 gene_biotype:protein_coding transcript_biotype:protein_coding gene_symbol:TBX3 description:T-box transcription factor 3 [Source:HGNC Symbol;Acc:HGNC:11602]
>ENSP00000392510.2 pep chromosome:GRCh38:11:36274223:36289449:-1 gene:ENSG00000110442.12 transcript:ENST00000452374.6 gene_biotype:protein_coding transcript_biotype:protein_coding gene_symbol:COMMD9 description:COMM domain containing 9 [Source:HGNC Symbol;Acc:HGNC:25014]
>ENSP00000440260.1 pep chromosome:GRCh38:11:113804928:113809175:-1 gene:ENSG00000048028.12 transcript:ENST00000538224.1 gene_biotype:protein_coding transcript_biotype:protein_coding gene_symbol:USP28 description:ubiquitin specific peptidase 28 [Source:HGNC Symbol;Acc:HGNC:12625]
>ENSP00000489147.3 pep chromosome:GRCh38:22:50674892:50733212:1 gene:ENSG00000251322.12 transcript:ENST00000262795.8 gene_biotype:protein_coding transcript_biotype:protein_coding gene_symbol:SHANK3 description:SH3 and multiple ankyrin repeat domains 3 [Source:HGNC Symbol;Acc:HGNC:14294]
>a615f1a95ea01e8999701b297af1e184
>dd1d3613798bb172df432862b7a83db9
>rev_13ff16aca624126d9a4835a48a1a064e
>rev_4b046db543d0bd359c850850305bac60
>rev_826d81b1ff05d1e99a9966d3f4bed7b1
>rev_ENSP00000346342.2 pep chromosome:GRCh38:22:29438583:29442455:1 gene:ENSG00000128250.5 transcript:ENST00000354373.2 gene_biotype:protein_coding transcript_biotype:protein_coding gene_symbol:RFPL1 description:ret finger protein like 1 [Source:HGNC Symbol;Acc:HGNC:9977]
>rev_ENSP00000413135.1 pep chromosome:GRCh38:19:44905791:44908944:1 gene:ENSG00000130203.10 transcript:ENST00000446996.5 gene_biotype:protein_coding transcript_biotype:protein_coding gene_symbol:APOE description:apolipoprotein E [Source:HGNC Symbol;Acc:HGNC:613]
>rev_ENSP00000458983.1 pep chromosome:GRCh38:17:6628573:6640662:-1 gene:ENSG00000198920.11 transcript:ENST00000571642.5 gene_biotype:protein_coding transcript_biotype:protein_coding gene_symbol:KIAA0753 description:KIAA0753 [Source:HGNC Symbol;Acc:HGNC:29110]
>rev_ENSP00000502121.1 pep chromosome:GRCh38:5:123344895:123423421:-1 gene:ENSG00000168944.17 transcript:ENST00000675814.1 gene_biotype:protein_coding transcript_biotype:nonsense_mediated_decay gene_symbol:CEP120 description:centrosomal protein 120 [Source:HGNC Symbol;Acc:HGNC:26690]
>rev_ba92728450ed8b4fe9014f527d19863e
>rev_f1b8b8dbfa68938f92c3b9e31b633204
>rev_sp|Q7KZI7-16|MARK2_HUMAN Isoform 16 of Serine/threonine-protein kinase MARK2 OS=Homo sapiens OX=9606 GN=MARK2
>sp|P48960-2|AGRE5_HUMAN Isoform 2 of Adhesion G protein-coupled receptor E5 OS=Homo sapiens OX=9606 GN=ADGRE5
>sp|W6CW81|PYDC5_HUMAN Pyrin domain-containing protein 5 OS=Homo sapiens OX=9606 GN=PYDC5 PE=1 SV=1
# FragPipe v23.0ui state cache
# Please edit the following path to point to the correct location.
# In Windows, please replace single '\' with '\\'
database.db-path=/home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/2026-02-04-decoys-contam-2025-11-27__14-31-59.f70db2649f5e8c3c1331271851956fd7.merged.fas.fas
crystalc.run-crystalc=false
database.decoy-tag=rev_
diann.channel-normalization-strategy=0
diann.cmd-opts=
diann.generate-msstats=true
diann.heavy=
diann.library=
diann.light=
diann.medium=
diann.min-site-prob=0.75
diann.mod-tag=
diann.q-value=0.01
diann.quantification-strategy=3
diann.quantification-strategy-2=2
diann.run-dia-nn=true
diann.run-dia-plex=false
diann.run-specific-protein-q-value=false
diann.unrelated-runs=false
diann.use-predicted-spectra=false
diatracer.corr-threshold=0.3
diatracer.delta-apex-im=0.01
diatracer.delta-apex-rt=3
diatracer.mass-defect-filter=true
diatracer.mass-defect-offset=0.1
diatracer.rf-max=500
diatracer.run-diatracer=false
diatracer.write-intermediate-files=false
diaumpire.AdjustFragIntensity=true
diaumpire.BoostComplementaryIon=false
diaumpire.CorrThreshold=0
diaumpire.DeltaApex=0.2
diaumpire.ExportPrecursorPeak=false
diaumpire.Q1=true
diaumpire.Q2=true
diaumpire.Q3=true
diaumpire.RFmax=500
diaumpire.RPmax=25
diaumpire.RTOverlap=0.3
diaumpire.SE.EstimateBG=false
diaumpire.SE.IsoPattern=0.3
diaumpire.SE.MS1PPM=10
diaumpire.SE.MS2PPM=20
diaumpire.SE.MS2SN=1.1
diaumpire.SE.MassDefectFilter=true
diaumpire.SE.MassDefectOffset=0.1
diaumpire.SE.NoMissedScan=1
diaumpire.SE.SN=1.1
diaumpire.run-diaumpire=false
fpop.coadaptr.fpop.fpop_masses=
fpop.coadaptr.fpop.run-fpop-coadaptr=false
fpop.fragpipe.fpop.fpop-tmt=false
fpop.fragpipe.fpop.label_control=
fpop.fragpipe.fpop.label_fpop=
fpop.fragpipe.fpop.region_size=1
fpop.fragpipe.fpop.run-fpop=false
fpop.fragpipe.fpop.subtract-control=false
fragpipe-config.bin-diann=/fragpipe_bin/fragpipe-23.0/fragpipe-23.0/tools/diann/1.8.2_beta_8/linux/diann-1.8.1.8
fragpipe-config.bin-python=python3
fragpipe-config.tools-folder=/home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940/tools_dir
freequant.mz-tol=10
freequant.rt-tol=0.4
freequant.run-freequant=false
ionquant.excludemods=
ionquant.formula=
ionquant.heavy=
ionquant.imtol=0.05
ionquant.ionfdr=0.01
ionquant.light=
ionquant.locprob=0.75
ionquant.maxlfq=1
ionquant.mbr=0
ionquant.mbrimtol=0.05
ionquant.mbrmincorr=0
ionquant.mbrrttol=1
ionquant.mbrtoprun=10
ionquant.medium=
ionquant.minfreq=0
ionquant.minions=1
ionquant.minisotopes=1
ionquant.minscans=3
ionquant.mztol=10
ionquant.normalization=1
ionquant.peptidefdr=1
ionquant.proteinfdr=1
ionquant.requantify=1
ionquant.rttol=0.4
ionquant.run-ionquant=true
ionquant.tp=0
ionquant.uniqueness=0
ionquant.use-labeling=false
ionquant.use-lfq=true
ionquant.writeindex=0
msbooster.find-best-im-model=false
msbooster.find-best-rt-model=false
msbooster.find-best-spectra-model=false
msbooster.fragmentation-type=0
msbooster.im-model=DIA-NN
msbooster.koina-url=
msbooster.predict-im=true
msbooster.predict-rt=true
msbooster.predict-spectra=true
msbooster.rt-model=DIA-NN
msbooster.run-msbooster=true
msbooster.spectra-model=DIA-NN
msbooster.spectral-library-path=
msfragger.Y_type_masses=
msfragger.activation_types=all
msfragger.allowed_missed_cleavage_1=1
msfragger.allowed_missed_cleavage_2=1
msfragger.analyzer_types=all
msfragger.calibrate_mass=2
msfragger.check_spectral_files=false
msfragger.clip_nTerm_M=true
msfragger.deisotope=1
msfragger.delta_mass_exclude_ranges=(-1.5,3.5)
msfragger.deneutralloss=1
msfragger.diagnostic_fragments=
msfragger.diagnostic_intensity_filter=0
msfragger.digest_max_length=50
msfragger.digest_min_length=7
msfragger.fragment_ion_series=b,y
msfragger.fragment_mass_tolerance=20
msfragger.fragment_mass_units=1
msfragger.group_variable=0
msfragger.intensity_transform=0
msfragger.ion_series_definitions=
msfragger.isotope_error=0/1/2
msfragger.labile_search_mode=off
msfragger.localize_delta_mass=false
msfragger.mass_diff_to_variable_mod=0
msfragger.mass_offsets=0
msfragger.mass_offsets_detailed=
msfragger.max_fragment_charge=2
msfragger.max_variable_mods_combinations=5000
msfragger.max_variable_mods_per_peptide=3
msfragger.min_fragments_modelling=2
msfragger.min_matched_fragments=4
msfragger.min_sequence_matches=2
msfragger.minimum_peaks=15
msfragger.minimum_ratio=0.00
msfragger.misc.fragger.clear-mz-hi=0
msfragger.misc.fragger.clear-mz-lo=0
msfragger.misc.fragger.digest-mass-hi=5000
msfragger.misc.fragger.digest-mass-lo=500
msfragger.misc.fragger.enzyme-dropdown-1=stricttrypsin
msfragger.misc.fragger.enzyme-dropdown-2=null
msfragger.misc.fragger.precursor-charge-hi=4
msfragger.misc.fragger.precursor-charge-lo=2
msfragger.misc.fragger.remove-precursor-range-hi=1.5
msfragger.misc.fragger.remove-precursor-range-lo=-1.5
msfragger.misc.slice-db=1
msfragger.num_enzyme_termini=2
msfragger.output_format=pepXML_pin
msfragger.output_max_expect=50
msfragger.output_report_topN=1
msfragger.output_report_topN_dda_plus=5
msfragger.output_report_topN_dia1=5
msfragger.override_charge=false
msfragger.precursor_mass_lower=-20
msfragger.precursor_mass_mode=selected
msfragger.precursor_mass_units=1
msfragger.precursor_mass_upper=20
msfragger.precursor_true_tolerance=20
msfragger.precursor_true_units=1
msfragger.remainder_fragment_masses=
msfragger.remove_precursor_peak=1
msfragger.report_alternative_proteins=true
msfragger.require_precursor=true
msfragger.restrict_deltamass_to=all
msfragger.reuse_dia_fragment_peaks=false
msfragger.run-msfragger=true
msfragger.search_enzyme_cut_1=KR
msfragger.search_enzyme_cut_2=
msfragger.search_enzyme_name_1=stricttrypsin
msfragger.search_enzyme_name_2=null
msfragger.search_enzyme_nocut_1=
msfragger.search_enzyme_nocut_2=
msfragger.search_enzyme_sense_1=C
msfragger.search_enzyme_sense_2=C
msfragger.table.fix-mods=0.0,C-Term Peptide,true,-1; 0.0,N-Term Peptide,true,-1; 0.0,C-Term Protein,true,-1; 0.0,N-Term Protein,true,-1; 0.0,G (glycine),true,-1; 0.0,A (alanine),true,-1; 0.0,S (serine),true,-1; 0.0,P (proline),true,-1; 0.0,V (valine),true,-1; 0.0,T (threonine),true,-1; 57.02146,C (cysteine),true,-1; 0.0,L (leucine),true,-1; 0.0,I (isoleucine),true,-1; 0.0,N (asparagine),true,-1; 0.0,D (aspartic acid),true,-1; 0.0,Q (glutamine),true,-1; 0.0,K (lysine),true,-1; 0.0,E (glutamic acid),true,-1; 0.0,M (methionine),true,-1; 0.0,H (histidine),true,-1; 0.0,F (phenylalanine),true,-1; 0.0,R (arginine),true,-1; 0.0,Y (tyrosine),true,-1; 0.0,W (tryptophan),true,-1; 0.0,B ,true,-1; 0.0,J,true,-1; 0.0,O,true,-1; 0.0,U,true,-1; 0.0,X,true,-1; 0.0,Z,true,-1
msfragger.table.var-mods=15.9949,M,true,1; 42.0106,[^,true,1; 79.96633,STY,false,3; -17.0265,nQnC,false,1; -18.0106,nE,false,1; 0.0,site_06,false,1; 0.0,site_07,false,1; 0.0,site_08,false,1; 0.0,site_09,false,1; 0.0,site_10,false,1; 0.0,site_11,false,1; 0.0,site_12,false,1; 0.0,site_13,false,1; 0.0,site_14,false,1; 0.0,site_15,false,1; 0.0,site_16,false,1
msfragger.track_zero_topN=0
msfragger.use_all_mods_in_first_search=false
msfragger.use_detailed_offsets=false
msfragger.use_topN_peaks=1000
msfragger.write_calibrated_mzml=false
msfragger.zero_bin_accept_expect=0
msfragger.zero_bin_mult_expect=1
opair.activation1=HCD
opair.activation2=ETD
opair.allowed_sites=
opair.filterOxonium=true
opair.glyco_db=
opair.max_glycans=4
opair.max_isotope_error=2
opair.min_isotope_error=0
opair.ms1_tol=20
opair.ms2_tol=20
opair.oxonium_filtering_file=
opair.oxonium_minimum_intensity=0.05
opair.reverse_scan_order=false
opair.run-opair=false
opair.single_scan_type=false
peptide-prophet.cmd-opts=--decoyprobs --ppm --accmass --nonparam --expectscore
peptide-prophet.combine-pepxml=false
peptide-prophet.run-peptide-prophet=false
percolator.cmd-opts=--no-terminate --post-processing-tdc --subset-max-train 500000
percolator.keep-tsv-files=false
percolator.min-prob=0.7
percolator.run-percolator=true
phi-report.dont-use-prot-proph-file=false
phi-report.filter=--picked --prot 0.01 --minPepLen 8
phi-report.pep-level-summary=false
phi-report.print-decoys=false
phi-report.prot-level-summary=false
phi-report.remove-contaminants=false
phi-report.run-report=true
protein-prophet.cmd-opts=--maxppmdiff 2000000 --minprob 0.5
protein-prophet.run-protein-prophet=true
ptmprophet.cmdline=
ptmprophet.override-defaults=false
ptmprophet.run-ptmprophet=false
ptmshepherd.adv_params=false
ptmshepherd.annotate_assigned_mods=false
ptmshepherd.annotation-common=false
ptmshepherd.annotation-custom=false
ptmshepherd.annotation-glyco=false
ptmshepherd.annotation-unimod=true
ptmshepherd.annotation_file=
ptmshepherd.annotation_tol=0.01
ptmshepherd.cap_y_ions=
ptmshepherd.decoy_type=1
ptmshepherd.diag_ions=
ptmshepherd.diagmine_diagMinFoldChange=3.0
ptmshepherd.diagmine_diagMinSpecDiff=25
ptmshepherd.diagmine_fragMinFoldChange=3.0
ptmshepherd.diagmine_fragMinPropensity=12.5
ptmshepherd.diagmine_fragMinSpecDiff=25
ptmshepherd.diagmine_minIonsPerSpec=2
ptmshepherd.diagmine_minPeps=25
ptmshepherd.diagmine_pepMinFoldChange=3.0
ptmshepherd.diagmine_pepMinSpecDiff=25
ptmshepherd.glyco_fdr=1.00
ptmshepherd.glyco_isotope_max=3
ptmshepherd.glyco_isotope_min=-1
ptmshepherd.glyco_ppm_tol=50
ptmshepherd.glycodatabase=
ptmshepherd.histo_smoothbins=2
ptmshepherd.iontype_a=false
ptmshepherd.iontype_b=true
ptmshepherd.iontype_c=true
ptmshepherd.iontype_x=false
ptmshepherd.iontype_y=true
ptmshepherd.iontype_z=true
ptmshepherd.localization_allowed_res=
ptmshepherd.n_glyco=true
ptmshepherd.normalization-psms=true
ptmshepherd.normalization-scans=false
ptmshepherd.output_extended=false
ptmshepherd.peakpicking_mass_units=0
ptmshepherd.peakpicking_minPsm=10
ptmshepherd.peakpicking_promRatio=0.3
ptmshepherd.peakpicking_width=0.002
ptmshepherd.precursor_mass_units=0
ptmshepherd.precursor_tol=0.01
ptmshepherd.print_decoys=false
ptmshepherd.print_full_glyco_params=false
ptmshepherd.prob_mass=0.5
ptmshepherd.remainder_masses=
ptmshepherd.remove_glycan_delta_mass=true
ptmshepherd.run-shepherd=false
ptmshepherd.run_diagextract_mode=false
ptmshepherd.run_diagmine_mode=false
ptmshepherd.run_glyco_mode=false
ptmshepherd.spectra_condPeaks=150
ptmshepherd.spectra_condRatio=0.0001
ptmshepherd.spectra_maxPrecursorCharge=4
ptmshepherd.spectra_maxfragcharge=2
ptmshepherd.spectra_ppmtol=20
ptmshepherd.use_msfragger_localization=true
ptmshepherd.varmod_masses=
quantitation.run-label-free-quant=false
run-psm-validation=true
run-validation-tab=true
saintexpress.cmd-opts=
saintexpress.max-replicates=10
saintexpress.run-saint-express=false
saintexpress.virtual-controls=100
skyline.run-skyline=false
skyline.skyline=false
skyline.skyline-custom=false
skyline.skyline-custom-path=
skyline.skyline-daily=true
skyline.skyline-fragment-tolerance=10
skyline.skyline-mods-mode=Default
skyline.skyline-precursor-tolerance=10
skyline.use-ssl=false
speclibgen.convert-pepxml=false
speclibgen.convert-psm=true
speclibgen.easypqp.extras.max_delta_ppm=15
speclibgen.easypqp.extras.max_delta_unimod=0.02
speclibgen.easypqp.extras.max_glycan_qval=1
speclibgen.easypqp.extras.rt_lowess_fraction=0
speclibgen.easypqp.fragment.a=false
speclibgen.easypqp.fragment.b=true
speclibgen.easypqp.fragment.c=false
speclibgen.easypqp.fragment.x=false
speclibgen.easypqp.fragment.y=true
speclibgen.easypqp.fragment.z=false
speclibgen.easypqp.im-cal=Automatic selection of a run as reference IM
speclibgen.easypqp.labile_mode=Regular (not glyco)
speclibgen.easypqp.neutral_loss=false
speclibgen.easypqp.rt-cal=noiRT
speclibgen.easypqp.select-file.text=
speclibgen.easypqp.select-im-file.text=
speclibgen.keep-intermediate-files=false
speclibgen.run-speclibgen=true
tab-run.delete_temp_files=false
tab-run.export_matched_fragments=false
tab-run.sub_mzml_prob_threshold=0.5
tab-run.write_sub_mzml=false
tmtintegrator.add_Ref=-1
tmtintegrator.aggregation_method=0
tmtintegrator.allow_overlabel=true
tmtintegrator.allow_unlabeled=true
tmtintegrator.best_psm=true
tmtintegrator.channel_num=TMT-6
tmtintegrator.extraction_tool=IonQuant
tmtintegrator.glyco_qval=-1
tmtintegrator.groupby=-1
tmtintegrator.log2transformed=true
tmtintegrator.max_pep_prob_thres=0
tmtintegrator.min_ntt=0
tmtintegrator.min_pep_prob=0.9
tmtintegrator.min_percent=0.05
tmtintegrator.min_purity=0.5
tmtintegrator.min_resolution=0
tmtintegrator.min_site_prob=-1
tmtintegrator.min_snr=0
tmtintegrator.mod_tag=none
tmtintegrator.ms1_int=true
tmtintegrator.outlier_removal=true
tmtintegrator.philosopher-msstats=false
tmtintegrator.print_RefInt=false
tmtintegrator.prot_exclude=none
tmtintegrator.prot_norm=0
tmtintegrator.psm_norm=false
tmtintegrator.quant_level=2
tmtintegrator.ref_d_tag=Pool
tmtintegrator.ref_tag=Bridge
tmtintegrator.run-tmtintegrator=false
tmtintegrator.tolerance=20
tmtintegrator.unique_gene=0
tmtintegrator.unique_pep=false
tmtintegrator.use_glycan_composition=false
workdir=/home/jliu/VandT_proteomics/DIA_Exploris_00/work/ce/c6d675e59d4485cf6531ed45e27940
workflow.input.data-type.im-ms=false
workflow.input.data-type.regular-ms=true
workflow.misc.save-sdrf=true
workflow.misc.sdrf-type=Default
workflow.ram=37
workflow.threads=35
<img width="1642" height="1428" alt="Image" src="https://github.com/user-attachments/assets/ec10767b-134b-424d-b018-7cc55e55e35d" />
<img width="1588" height="1002" alt="Image" src="https://github.com/user-attachments/assets/425b98c9-db66-45b1-adf7-c775485a254e" />