DIA-NN 1.8.1 (Data-Independent Acquisition by Neural Networks) Compiled on Apr 14 2022 15:31:19 Current date and time: Wed Feb 26 12:08:27 2025 CPU: GenuineIntel Intel(R) Xeon(R) W-2235 CPU @ 3.80GHz SIMD instructions: AVX AVX2 AVX512CD AVX512F FMA SSE4.1 SSE4.2 Logical CPU cores: 12 diann.exe --f D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_1.raw --f D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_2.raw --f D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_3.raw --f D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_4.raw --f D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_5.raw --f D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_6.raw --f D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_7.raw --f D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_8.raw --f D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_9.raw --f D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_10.raw --f D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_11.raw --lib --threads 6 --verbose 1 --out D:\data\DIA-NN\250220_FH_YS1\250220_FH_YS1.tsv --qvalue 0.01 --matrices --predictor --fasta D:\data\FASTA\original\uniprotkb_taxonomy_id_31033_2025_02_04_canonical_isoform.fasta --fasta-search --min-fr-mz 145 --max-fr-mz 1450 --met-excision --cut K*,R*,!*P --missed-cleavages 1 --min-pep-len 7 --max-pep-len 30 --min-pr-mz 460 --max-pr-mz 860 --min-pr-charge 2 --max-pr-charge 4 --unimod4 --var-mods 1 --var-mod UniMod:35,15.994915,M --var-mod UniMod:1,42.010565,*n --monitor-mod UniMod:1 --window 8 --no-isotopes --smart-profiling --pg-level 1 --peak-center --no-ifs-removal Thread number set to 6 Output will be filtered at 0.01 FDR Precursor/protein x samples expression level matrices will be saved along with the main report Deep learning will be used to generate a new in silico spectral library from peptides provided Library-free search enabled Min fragment m/z set to 145 Max fragment m/z set to 1450 N-terminal methionine excision enabled In silico digest will involve cuts at K*,R* But excluding cuts at *P Maximum number of missed cleavages set to 1 Min peptide length set to 7 Max peptide length set to 30 Min precursor m/z set to 460 Max precursor m/z set to 860 Min precursor charge set to 2 Max precursor charge set to 4 Cysteine carbamidomethylation enabled as a fixed modification Maximum number of variable modifications set to 1 Modification UniMod:35 with mass delta 15.9949 at M will be considered as variable Modification UniMod:1 with mass delta 42.0106 at *n will be considered as variable Scan window radius set to 8 Isotopologue chromatograms will not be used When generating a spectral library, in silico predicted spectra will be retained if deemed more reliable than experimental ones Implicit protein grouping: protein names; this determines which peptides are considered 'proteotypic' and thus affects protein FDR calculation Fixed-width center of each elution peak will be used for quantification Interference removal from fragment elution curves disabled DIA-NN will optimise the mass accuracy automatically using the first run in the experiment. This is useful primarily for quick initial analyses, when it is not yet known which mass accuracy setting works best for a particular acquisition scheme. Exclusion of fragments shared between heavy and light peptides from quantification is not supported in FASTA digest mode - disabled; to enable, generate an in silico predicted spectral library and analyse with this library The following variable modifications will be scored: UniMod:1 11 files will be processed [0:00] Loading FASTA D:\data\FASTA\original\uniprotkb_taxonomy_id_31033_2025_02_04_canonical_isoform.fasta [0:09] Processing FASTA [0:18] Assembling elution groups [0:25] 2575737 precursors generated [0:26] Gene names missing for some isoforms [0:26] Library contains 52309 proteins, and 14284 genes [0:26] [0:31] [17:14] [21:38] [21:48] [21:50] Saving the library to lib.predicted.speclib [21:56] Initialising library [21:57] File #1/11 [21:57] Loading run D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_1.raw [22:52] 2575737 library precursors are potentially detectable [22:53] Processing... [23:33] RT window set to 4.00098 [23:33] Recommended MS1 mass accuracy setting: 5.14341 ppm [24:21] Optimised mass accuracy: 12.7928 ppm [27:58] Removing low confidence identifications [27:58] Searching PTM decoys [28:02] Removing interfering precursors [28:09] Training neural networks: 136149 targets, 88623 decoys [28:22] Number of IDs at 0.01 FDR: 70918 [28:23] Calculating protein q-values [28:23] Number of proteins identified at 1% FDR: 4380 (precursor-level), 3953 (protein-level) (inference performed using proteotypic peptides only) [28:24] Quantification [28:25] Precursors with monitored PTMs at 1% FDR: 429 out of 526 [28:25] Unmodified precursors with monitored PTM sites at 1% FDR: 151 out of 200 [28:26] Quantification information saved to D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_1.raw.quant. [28:26] File #2/11 [28:26] Loading run D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_2.raw [29:52] 2575737 library precursors are potentially detectable [29:53] Processing... [30:41] RT window set to 4.15809 [30:42] Recommended MS1 mass accuracy setting: 5.03272 ppm [34:36] Removing low confidence identifications [34:37] Searching PTM decoys [34:41] Removing interfering precursors [34:47] Training neural networks: 136603 targets, 85838 decoys [35:00] Number of IDs at 0.01 FDR: 67104 [35:01] Calculating protein q-values [35:02] Number of proteins identified at 1% FDR: 4213 (precursor-level), 3836 (protein-level) (inference performed using proteotypic peptides only) [35:02] Quantification [35:03] Precursors with monitored PTMs at 1% FDR: 349 out of 454 [35:03] Unmodified precursors with monitored PTM sites at 1% FDR: 134 out of 196 [35:03] Quantification information saved to D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_2.raw.quant. [35:04] File #3/11 [35:04] Loading run D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_3.raw [35:41] 2575737 library precursors are potentially detectable [35:41] Processing... [36:27] RT window set to 4.02744 [36:27] Recommended MS1 mass accuracy setting: 5.15372 ppm [40:00] Removing low confidence identifications [40:00] Searching PTM decoys [40:04] Removing interfering precursors [40:11] Training neural networks: 135690 targets, 86939 decoys [40:24] Number of IDs at 0.01 FDR: 68446 [40:25] Calculating protein q-values [40:25] Number of proteins identified at 1% FDR: 4318 (precursor-level), 3874 (protein-level) (inference performed using proteotypic peptides only) [40:25] Quantification [40:26] Precursors with monitored PTMs at 1% FDR: 410 out of 562 [40:26] Unmodified precursors with monitored PTM sites at 1% FDR: 131 out of 211 [40:27] Quantification information saved to D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_3.raw.quant. [40:28] File #4/11 [40:28] Loading run D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_4.raw [41:07] 2575737 library precursors are potentially detectable [41:07] Processing... [41:53] RT window set to 4.4612 [41:54] Recommended MS1 mass accuracy setting: 5.41024 ppm [45:51] Removing low confidence identifications [45:51] Searching PTM decoys [45:55] Removing interfering precursors [46:02] Training neural networks: 145474 targets, 93728 decoys [46:16] Number of IDs at 0.01 FDR: 74609 [46:17] Calculating protein q-values [46:18] Number of proteins identified at 1% FDR: 4303 (precursor-level), 3825 (protein-level) (inference performed using proteotypic peptides only) [46:18] Quantification [46:19] Precursors with monitored PTMs at 1% FDR: 415 out of 483 [46:19] Unmodified precursors with monitored PTM sites at 1% FDR: 167 out of 210 [46:20] Quantification information saved to D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_4.raw.quant. [46:20] File #5/11 [46:20] Loading run D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_5.raw [46:57] 2575737 library precursors are potentially detectable [46:57] Processing... [47:43] RT window set to 4.26721 [47:43] Recommended MS1 mass accuracy setting: 5.61987 ppm [51:31] Removing low confidence identifications [51:31] Searching PTM decoys [51:36] Removing interfering precursors [51:42] Training neural networks: 143735 targets, 93116 decoys [51:57] Number of IDs at 0.01 FDR: 72340 [51:58] Calculating protein q-values [51:58] Number of proteins identified at 1% FDR: 4191 (precursor-level), 3771 (protein-level) (inference performed using proteotypic peptides only) [51:58] Quantification [51:59] Precursors with monitored PTMs at 1% FDR: 529 out of 614 [51:59] Unmodified precursors with monitored PTM sites at 1% FDR: 191 out of 232 [52:00] Quantification information saved to D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_5.raw.quant. [52:01] File #6/11 [52:01] Loading run D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_6.raw [52:40] 2575737 library precursors are potentially detectable [52:40] Processing... [53:27] RT window set to 4.14068 [53:27] Recommended MS1 mass accuracy setting: 5.11005 ppm [57:09] Removing low confidence identifications [57:09] Searching PTM decoys [57:13] Removing interfering precursors [57:19] Training neural networks: 133392 targets, 84639 decoys [57:32] Number of IDs at 0.01 FDR: 67554 [57:33] Calculating protein q-values [57:33] Number of proteins identified at 1% FDR: 4361 (precursor-level), 3960 (protein-level) (inference performed using proteotypic peptides only) [57:33] Quantification [57:35] Precursors with monitored PTMs at 1% FDR: 452 out of 495 [57:35] Unmodified precursors with monitored PTM sites at 1% FDR: 178 out of 201 [57:35] Quantification information saved to D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_6.raw.quant. [57:36] File #7/11 [57:36] Loading run D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_7.raw [58:15] 2575737 library precursors are potentially detectable [58:16] Processing... [58:55] RT window set to 4.10761 [58:56] Recommended MS1 mass accuracy setting: 5.07498 ppm [62:40] Removing low confidence identifications [62:40] Searching PTM decoys [62:44] Removing interfering precursors [62:51] Training neural networks: 154464 targets, 99032 decoys [63:06] Number of IDs at 0.01 FDR: 77582 [63:07] Calculating protein q-values [63:07] Number of proteins identified at 1% FDR: 4746 (precursor-level), 4186 (protein-level) (inference performed using proteotypic peptides only) [63:08] Quantification [63:09] Precursors with monitored PTMs at 1% FDR: 423 out of 491 [63:09] Unmodified precursors with monitored PTM sites at 1% FDR: 163 out of 215 [63:10] Quantification information saved to D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_7.raw.quant. [63:10] File #8/11 [63:10] Loading run D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_8.raw [63:48] 2575737 library precursors are potentially detectable [63:48] Processing... [64:27] RT window set to 4.12526 [64:28] Recommended MS1 mass accuracy setting: 5.84635 ppm [68:06] Removing low confidence identifications [68:07] Searching PTM decoys [68:11] Removing interfering precursors [68:17] Training neural networks: 140574 targets, 89777 decoys [68:31] Number of IDs at 0.01 FDR: 71086 [68:32] Calculating protein q-values [68:32] Number of proteins identified at 1% FDR: 4416 (precursor-level), 3938 (protein-level) (inference performed using proteotypic peptides only) [68:32] Quantification [68:34] Precursors with monitored PTMs at 1% FDR: 439 out of 514 [68:34] Unmodified precursors with monitored PTM sites at 1% FDR: 165 out of 220 [68:34] Quantification information saved to D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_8.raw.quant. [68:35] File #9/11 [68:35] Loading run D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_9.raw [69:05] 2575737 library precursors are potentially detectable [69:06] Processing... [69:51] RT window set to 4.14011 [69:52] Recommended MS1 mass accuracy setting: 5.3089 ppm [73:18] Removing low confidence identifications [73:18] Searching PTM decoys [73:22] Removing interfering precursors [73:29] Training neural networks: 128473 targets, 82186 decoys [73:41] Number of IDs at 0.01 FDR: 64932 [73:42] Calculating protein q-values [73:42] Number of proteins identified at 1% FDR: 4264 (precursor-level), 3880 (protein-level) (inference performed using proteotypic peptides only) [73:42] Quantification [73:43] Precursors with monitored PTMs at 1% FDR: 425 out of 482 [73:43] Unmodified precursors with monitored PTM sites at 1% FDR: 166 out of 197 [73:44] Quantification information saved to D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_9.raw.quant. [73:45] File #10/11 [73:45] Loading run D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_10.raw [74:29] 2575737 library precursors are potentially detectable [74:29] Processing... [75:16] RT window set to 4.34826 [75:16] Recommended MS1 mass accuracy setting: 5.96639 ppm [79:21] Removing low confidence identifications [79:21] Searching PTM decoys [79:26] Removing interfering precursors [79:32] Training neural networks: 145128 targets, 92488 decoys [79:46] Number of IDs at 0.01 FDR: 72357 [79:47] Calculating protein q-values [79:48] Number of proteins identified at 1% FDR: 4529 (precursor-level), 4081 (protein-level) (inference performed using proteotypic peptides only) [79:48] Quantification [79:49] Precursors with monitored PTMs at 1% FDR: 518 out of 563 [79:49] Unmodified precursors with monitored PTM sites at 1% FDR: 194 out of 218 [79:50] Quantification information saved to D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_10.raw.quant. [79:51] File #11/11 [79:51] Loading run D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_11.raw [80:20] 2575737 library precursors are potentially detectable [80:20] Processing... [81:03] RT window set to 4.26869 [81:04] Recommended MS1 mass accuracy setting: 4.79475 ppm [84:34] Removing low confidence identifications [84:35] Searching PTM decoys [84:39] Removing interfering precursors [84:45] Training neural networks: 134703 targets, 87923 decoys [84:59] Number of IDs at 0.01 FDR: 69470 [85:00] Calculating protein q-values [85:00] Number of proteins identified at 1% FDR: 4265 (precursor-level), 3844 (protein-level) (inference performed using proteotypic peptides only) [85:00] Quantification [85:02] Precursors with monitored PTMs at 1% FDR: 563 out of 634 [85:02] Unmodified precursors with monitored PTM sites at 1% FDR: 197 out of 249 [85:02] Quantification information saved to D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_11.raw.quant. [85:03] Cross-run analysis [85:03] Reading quantification information: 11 files [85:06] Quantifying peptides [85:13] Assembling protein groups [85:17] Quantifying proteins [85:18] Calculating q-values for protein and gene groups [85:19] Calculating global q-values for protein and gene groups [85:19] Writing report [86:01] Report saved to D:\data\DIA-NN\250220_FH_YS1\250220_FH_YS1.tsv. [86:01] Saving precursor levels matrix [86:02] Precursor levels matrix (1% precursor and protein group FDR) saved to D:\data\DIA-NN\250220_FH_YS1\250220_FH_YS1.pr_matrix.tsv. [86:02] Saving protein group levels matrix [86:02] Protein group levels matrix (1% precursor FDR and protein group FDR) saved to D:\data\DIA-NN\250220_FH_YS1\250220_FH_YS1.pg_matrix.tsv. [86:02] Saving gene group levels matrix [86:02] Gene groups levels matrix (1% precursor FDR and protein group FDR) saved to D:\data\DIA-NN\250220_FH_YS1\250220_FH_YS1.gg_matrix.tsv. [86:02] Saving unique genes levels matrix [86:03] Unique genes levels matrix (1% precursor FDR and protein group FDR) saved to D:\data\DIA-NN\250220_FH_YS1\250220_FH_YS1.unique_genes_matrix.tsv. [86:03] Stats report saved to D:\data\DIA-NN\250220_FH_YS1\250220_FH_YS1.stats.tsv [86:03] Generating spectral library: [86:03] 108607 precursors passing the FDR threshold are to be extracted [86:03] Loading run D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_1.raw [86:38] 2575737 library precursors are potentially detectable [86:39] 7374 spectra added to the library [86:40] Loading run D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_2.raw [87:22] 2575737 library precursors are potentially detectable [87:23] 5790 spectra added to the library [87:24] Loading run D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_3.raw [87:59] 2575737 library precursors are potentially detectable [87:59] 2708 spectra added to the library [88:00] Loading run D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_4.raw [88:39] 2575737 library precursors are potentially detectable [88:40] 13148 spectra added to the library [88:41] Loading run D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_5.raw [89:16] 2575737 library precursors are potentially detectable [89:17] 11119 spectra added to the library [89:18] Loading run D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_6.raw [89:57] 2575737 library precursors are potentially detectable [89:58] 3735 spectra added to the library [89:58] Loading run D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_7.raw [90:18] 2575737 library precursors are potentially detectable [90:19] 9940 spectra added to the library [90:20] Loading run D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_8.raw [90:38] 2575737 library precursors are potentially detectable [90:39] 7222 spectra added to the library [90:40] Loading run D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_9.raw [90:57] 2575737 library precursors are potentially detectable [90:58] 3873 spectra added to the library [90:58] Loading run D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_10.raw [91:19] 2575737 library precursors are potentially detectable [91:19] 4504 spectra added to the library [91:20] Loading run D:\data\Rawdata\250220_FH_YS1\OE_250220_FH_YS1_11.raw [91:37] 2575737 library precursors are potentially detectable [91:39] 16886 spectra added to the library [91:39] Saving spectral library to lib.tsv [91:53] 108607 precursors saved [91:53] Loading the generated library and saving it in the .speclib format [91:53] Loading spectral library lib.tsv [91:58] Spectral library loaded: 31744 protein isoforms, 20008 protein groups and 108607 precursors in 96749 elution groups. [91:58] Loading protein annotations from FASTA D:\data\FASTA\original\uniprotkb_taxonomy_id_31033_2025_02_04_canonical_isoform.fasta [92:00] Gene names missing for some isoforms [92:00] Library contains 31740 proteins, and 7986 genes [92:00] Saving the library to lib.tsv.speclib Finished