Detecting decoy prefix in 2_site_library_23_02_08.fasta. No common prefixes were found. No common suffixes were found. No decoy prefix detected, but an existing decoy database seems to exist at decoy_2_site_library_23_02_08.fasta. Retrying the decoy prefix calculation on that. Detecting decoy prefix in decoy_2_site_library_23_02_08.fasta. Some common prefixes were found: DECOY: 49.9% DEC: 49.9% Using DECOY as the most likely decoy prefix. MSFragger version MSFragger-3.4 Batmass-IO version 1.23.6 timsdata library version timsdata-2-8-7-1 (c) University of Michigan RawFileReader reading tool. Copyright (c) 2016 by Thermo Fisher Scientific, Inc. All rights reserved. System OS: Windows 10, Architecture: AMD64 Java Info: 17.0.1, OpenJDK 64-Bit Server VM, Oracle Corporation JVM started with 16 GB memory Checking database... Checking spectral files... 09:19:39:149 [pool-1-thread-7] INFO PlatformDependent - Your platform does not provide complete low-level API for accessing direct buffers reliably. Unless explicitly requested, heap buffer will always be preferred to avoid potential system instability. C:\Users\Owner\Desktop\Pentelute\LCMS_Data\314_Orbi_Start_22_11_21\314_23_01_20\314_23_01_20_1_JSA482.raw: Scans = 8422 C:\Users\Owner\Desktop\Pentelute\LCMS_Data\314_Orbi_Start_22_11_21\314_23_01_20\314_23_01_20_3_JSA482.raw: Scans = 9239 C:\Users\Owner\Desktop\Pentelute\LCMS_Data\314_Orbi_Start_22_11_21\314_23_01_20\314_23_01_20_2_JSA482.raw: Scans = 9246 C:\Users\Owner\Desktop\Pentelute\LCMS_Data\314_Orbi_Start_22_11_21\314_23_01_20\314_23_01_20_1_JSA485.raw: Scans = 10429 C:\Users\Owner\Desktop\Pentelute\LCMS_Data\314_Orbi_Start_22_11_21\314_23_01_20\314_23_01_20_3_JSA485.raw: Scans = 10623 C:\Users\Owner\Desktop\Pentelute\LCMS_Data\314_Orbi_Start_22_11_21\314_23_01_20\314_23_01_20_2_JSA485.raw: Scans = 10225 ************************************MAIN SEARCH************************************ Checking database... Parameters: num_threads = 12 database_name = C:\Users\Owner\Desktop\Pentelute\LCMS_Data\314_Orbi_Start_22_11_21\314_23_01_20\decoy_2_site_library_23_02_08.fasta decoy_prefix = DECOY precursor_mass_lower = -20.0 precursor_mass_upper = 20.0 precursor_mass_units = 1 data_type = 0 precursor_true_tolerance = 20.0 precursor_true_units = 1 fragment_mass_tolerance = 20.0 fragment_mass_units = 1 calibrate_mass = 0 use_all_mods_in_first_search = false write_calibrated_mgf = 0 isotope_error = 0/1/2/3 mass_offsets = 0 labile_search_mode = OFF restrict_deltamass_to = all precursor_mass_mode = SELECTED localize_delta_mass = false delta_mass_exclude_ranges = (-1.5,3.5) fragment_ion_series = b,y ion_series_definitions = search_enzyme_name = Met search_enzyme_sense_1 = C search_enzyme_cut_1 = M search_enzyme_nocut_1 = allowed_missed_cleavage_1 = 0 num_enzyme_termini = 2 clip_nTerm_M = true allow_multiple_variable_mods_on_residue = false max_variable_mods_per_peptide = 2 max_variable_mods_combinations = 5500 output_format = pepxml_pin output_report_topN = 1 output_max_expect = 50.0 report_alternative_proteins = true override_charge = false precursor_charge_low = 1 precursor_charge_high = 4 digest_min_length = 5 digest_max_length = 60 digest_mass_range_low = 200.0 digest_mass_range_high = 5000.0 max_fragment_charge = 2 deisotope = 1 deneutralloss = true track_zero_topN = 0 zero_bin_accept_expect = 0.0 zero_bin_mult_expect = 1.0 add_topN_complementary = 0 minimum_peaks = 15 use_topN_peaks = 50 minIonsScoring = 2 min_matched_fragments = 4 minimum_ratio = 0.01 intensity_transform = 0 remove_precursor_peak = 1 remove_precursor_range = -1.500000,1.500000 clear_mz_range_low = 0.0 clear_mz_range_high = 0.0 excluded_scan_list_file = mass_diff_to_variable_mod = 0 min_sequence_matches = 2 check_spectral_files = true variable_mod_01 = -0.984016 c^ 2 add_C_cysteine = 0 add_Cterm_peptide = 0.0 add_Cterm_protein = 0.0 add_Nterm_peptide = 0.0 add_Nterm_protein = 0.0 Number of unique peptides of length 14: 576 In total 576 peptides. Generated 1152 modified peptides. Number of peptides with more than 5500 modification patterns: 0 Selected fragment index width 0.10 Da. 29952 fragments to be searched in 1 slices (0.00 GB total) Operating on slice 1 of 1: Fragment index slice generated in 0.02 s 001. 314_23_01_20_1_JSA482.raw 16.3 s | deisotoping 0.3 s [progress: 7666/7666 (100%) - 23956 spectra/s] 0.3s | postprocessing 0.3 s 002. 314_23_01_20_1_JSA485.raw 12.6 s | deisotoping 0.3 s [progress: 10077/10077 (100%) - 42519 spectra/s] 0.2s | postprocessing 0.3 s 003. 314_23_01_20_2_JSA482.raw 12.1 s | deisotoping 0.0 s [progress: 8644/8644 (100%) - 77179 spectra/s] 0.1s | postprocessing 0.2 s 004. 314_23_01_20_2_JSA485.raw 12.2 s | deisotoping 0.2 s [progress: 9874/9874 (100%) - 86614 spectra/s] 0.1s | postprocessing 0.2 s 005. 314_23_01_20_3_JSA482.raw 12.5 s | deisotoping 0.0 s [progress: 8648/8648 (100%) - 81585 spectra/s] 0.1s | postprocessing 0.1 s 006. 314_23_01_20_3_JSA485.raw 12.2 s | deisotoping 0.0 s [progress: 10342/10342 (100%) - 92339 spectra/s] 0.1s | postprocessing 0.1 s ***************************MAIN SEARCH DONE IN 1.358 MIN*************************** *******************************TOTAL TIME 1.487 MIN******************************** WARNING: The output directory 'C:\Users\Owner\Desktop\Pentelute\LCMS_Data\314_Orbi_Start_22_11_21\314_23_01_20\crux-output' already exists. Existing files will be overwritten. WARNING: The file 'C:\Users\Owner\Desktop\Pentelute\LCMS_Data\314_Orbi_Start_22_11_21\314_23_01_20\crux-output/percolator.log.txt' already exists and will be overwritten. INFO: CPU: DESKTOP-J8TBBGG INFO: Crux version: 4.1 INFO: Wed 02/08/2023 INFO: Beginning percolator. INFO: Percolator version 3.05.nightly-137-e806a0c5, Build Date Oct 8 2021 22:15:04 INFO: Copyright (c) 2006-9 University of Washington. All rights reserved. INFO: Written by Lukas Käll (lukall@u.washington.edu) in the INFO: Department of Genome Sciences at the University of Washington. INFO: Issued command: INFO: percolator --results-peptides C:\Users\Owner\Desktop\Pentelute\LCMS_Data\314_Orbi_Start_22_11_21\314_23_01_20\crux-output/percolator.target.peptides.txt --decoy-results-peptides C:\Users\Owner\Desktop\Pentelute\LCMS_Data\314_Orbi_Start_22_11_21\314_23_01_20\crux-output/percolator.decoy.peptides.txt --results-psms C:\Users\Owner\Desktop\Pentelute\LCMS_Data\314_Orbi_Start_22_11_21\314_23_01_20\crux-output/percolator.target.psms.txt --decoy-results-psms C:\Users\Owner\Desktop\Pentelute\LCMS_Data\314_Orbi_Start_22_11_21\314_23_01_20\crux-output/percolator.decoy.psms.txt --verbose 2 --protein-decoy-pattern DECOY --seed 1 --subset-max-train 0 --trainFDR 0.01 --testFDR 0.05 --maxiter 10 --search-input auto --no-schema-validation --protein-enzyme trypsin --post-processing-tdc C:\Users\Owner\Desktop\Pentelute\LCMS_Data\314_Orbi_Start_22_11_21\314_23_01_20\314_23_01_20_1_JSA482.fixed.pin INFO: Started Wed Feb 8 09:21:09 2023 INFO: Hyperparameters: selectionFdr=0.01, Cpos=0, Cneg=0, maxNiter=10 INFO: Reading tab-delimited input from datafile C:\Users\Owner\Desktop\Pentelute\LCMS_Data\314_Orbi_Start_22_11_21\314_23_01_20\314_23_01_20_1_JSA482.fixed.pin INFO: Features: INFO: retentiontime rank abs_ppm isotope_errors log10_evalue hyperscore delta_hyperscore matched_ion_num complementary_ions ion_series weighted_average_abs_fragment_ppm peptide_length ntt nmc Charge1 Charge2 Charge3 Charge4 Charge5 Charge6 Charge7 INFO: Found 1452 PSMs INFO: Concatenated search input detected and --post-processing-tdc flag set. Applying target-decoy competition on Percolator scores. INFO: Train/test set contains 1446 positives and 6 negatives, size ratio=241 and pi0=1 INFO: Warning : the number of negative samples read is too small to perform a correct classification. INFO: INFO: Selecting Cpos by cross-validation. INFO: Selecting Cneg by cross-validation. INFO: Split 1: Selected feature 1 as initial direction. Could separate 964 training set positives with q<0.01 in that direction. INFO: Split 2: Selected feature 1 as initial direction. Could separate 964 training set positives with q<0.01 in that direction. INFO: Split 3: Selected feature 1 as initial direction. Could separate 964 training set positives with q<0.01 in that direction. INFO: Found 1446 test set positives with q<0.05 in initial direction INFO: Reading in data and feature calculation took 0.0200 cpu seconds or 0 seconds wall clock time. INFO: ---Training with Cpos selected by cross validation, Cneg selected by cross validation, initial_fdr=0.01, fdr=0.01 INFO: Iteration 1: Estimated 1446 PSMs with q<0.05 INFO: Iteration 2: Estimated 1446 PSMs with q<0.05 INFO: Iteration 3: Estimated 1446 PSMs with q<0.05 INFO: Iteration 4: Estimated 1446 PSMs with q<0.05 INFO: Iteration 5: Estimated 1446 PSMs with q<0.05 INFO: Iteration 6: Estimated 1446 PSMs with q<0.05 INFO: Iteration 7: Estimated 1446 PSMs with q<0.05 INFO: Iteration 8: Estimated 1446 PSMs with q<0.05 INFO: Iteration 9: Estimated 1446 PSMs with q<0.05 INFO: Iteration 10: Estimated 1446 PSMs with q<0.05 INFO: Learned normalized SVM weights for the 3 cross-validation splits: INFO: Split1 Split2 Split3 FeatureName INFO: 0.0958 0.2655 0.2696 retentiontime INFO: 0.0000 0.0000 0.0000 rank INFO: 0.0446 -0.1380 0.0823 abs_ppm INFO: 0.2980 -0.2672 -0.3288 isotope_errors INFO: -0.3641 -0.3398 -1.2171 log10_evalue INFO: 1.1681 2.1019 0.7024 hyperscore INFO: 0.3564 0.4118 0.3437 delta_hyperscore INFO: -0.3022 -0.9225 -0.5137 matched_ion_num INFO: 0.4287 0.4017 0.7068 complementary_ions INFO: -0.9929 -0.5768 -0.1683 ion_series INFO: -0.0555 -0.5604 -0.5191 weighted_average_abs_fragment_ppm INFO: 0.0000 0.0000 0.0000 peptide_length INFO: 0.0000 0.0000 0.0000 ntt INFO: 0.0000 0.0000 0.0000 nmc INFO: 0.0000 0.0000 0.0000 Charge1 INFO: 0.0000 0.0000 0.0000 Charge2 INFO: -0.1592 0.0934 -0.0235 Charge3 INFO: -0.1090 0.0870 -0.0324 Charge4 INFO: 0.2572 -0.1769 0.0564 Charge5 INFO: 0.0000 0.0000 0.0000 Charge6 INFO: 0.0000 0.0000 0.0000 Charge7 INFO: 2.1218 2.2190 2.5343 m0 INFO: Found 1446 test set PSMs with q<0.05. INFO: Selected best-scoring PSM per scan+expMass (target-decoy competition): 1446 target PSMs and 6 decoy PSMs. INFO: Multiple instantiations of Normalizer INFO: Tossing out "redundant" PSMs keeping only the best scoring PSM for each unique peptide. INFO: Calculating q values. INFO: Final list yields 160 target peptides with q<0.05. INFO: Calculating posterior error probabilities (PEPs). INFO: Processing took 5.0380 cpu seconds or 5 seconds wall clock time. INFO: Multiple instantiations of Normalizer WARNING: The file 'C:\Users\Owner\Desktop\Pentelute\LCMS_Data\314_Orbi_Start_22_11_21\314_23_01_20\crux-output/percolator.target.pep.xml' already exists and will be overwritten. FATAL: An exception occurred: 'c' is not a valid amino acid Search canceled.