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Nesvilab / FragPipe

A cross-platform proteomics data analysis suite
http://fragpipe.nesvilab.org
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Problem running software - keeps halting #120

Closed szackloo closed 5 years ago

szackloo commented 5 years ago

Describe the problem - seems to be a memory issue - have 16 Gb

System info

You can find that printed on the Config tab.

Describe your experiment

Extremely simple - purified protein binary complex (two - five proteins total) and digested by trypsin.

...

Input data files

mzml files

Sequence database

human
40000 entries

Attach fragger.params file

** _database_name = C:\Users\Suzanne Ackloo\Documents\Fasta files\Fasta files\2019-06-17-td-rev-UP000005640.fas num_threads = 4 # 0=poll CPU to set num threads; else specify num threads directly (max 64)

precursor_mass_lower = -50 precursor_mass_upper = 50 precursor_mass_units = 1 # 0=Daltons, 1=ppm precursor_true_tolerance = 30 precursor_true_units = 1 # 0=Daltons, 1=ppm fragment_mass_tolerance = 100 fragment_mass_units = 1 # 0=Daltons, 1=ppm calibrate_mass = 2 # 0=Off, 1=On, 2=On and find optimal parameters decoyprefix = rev

isotope_error = -1/0/1/2 # 0=off, -1/0/1/2/3 (standard C13 error) mass_offsets = 0 # allow for additional precursor mass window shifts. Multiplexed with isotope_error. mass_offsets = 0/79.966 can be used as a restricted ‘open’ search that looks for unmodified and phosphorylated peptides (on any residue) precursor_mass_mode = selected

localize_delta_mass = 0 delta_mass_exclude_ranges = (-1.5,3.5) fragment_ion_series = b,y

search_enzyme_name = Trypsin search_enzyme_cutafter = KR search_enzyme_butnotafter = P

num_enzyme_termini = 2 # 2 for enzymatic, 1 for semi-enzymatic, 0 for nonspecific digestion allowed_missed_cleavage = 2 # maximum value is 5

clip_nTerm_M = 0

maximum of 7 mods - amino acid codes, * for any amino acid, [ and ] specifies protein termini, n and c specifies peptide termini

variable_mod_01 = 15.99490 M variable_mod_02 = 42.01060 [^ variable_mod_03 = 79.96633 STY

variable_mod_04 = -17.02650 nQnC

variable_mod_05 = -18.01060 nE

variable_mod_06 = 0.00000 site_06

variable_mod_07 = 0.00000 site_07

allow_multiple_variable_mods_on_residue = 0 # static mods are not considered max_variable_mods_per_mod = 3 # maximum of 5 max_variable_mods_combinations = 5000 # maximum of 65534, limits number of modified peptides generated from sequence

output_file_extension = pepXML output_format = pepXML output_report_topN = 1 output_max_expect = 50 report_alternative_proteins = 1 # 0=no, 1=yes

precursor_charge = 2 4 # precursor charge range to analyze; does not override any existing charge; 0 as 1st entry ignores parameter override_charge = 0 # 0=no, 1=yes to override existing precursor charge states with precursor_charge parameter

digest_min_length = 7 digest_max_length = 50 digest_mass_range = 200.0 5000.0 # MH+ peptide mass range to analyze max_fragment_charge = 3 # set maximum fragment charge state to analyze (allowed max 5)

open search parameters

track_zero_topN = 0 # in addition to topN results, keep track of top results in zero bin zero_bin_accept_expect = 0.00 # boost top zero bin entry to top if it has expect under 0.01 - set to 0 to disable zero_bin_mult_expect = 1.00 # disabled if above passes - multiply expect of zero bin for ordering purposes (does not affect reported expect) add_topN_complementary = 0

spectral processing

minimum_peaks = 15 # required minimum number of peaks in spectrum to search (default 10) use_topN_peaks = 100 min_fragments_modelling = 3 min_matched_fragments = 4 minimum_ratio = 0.01 # filter peaks below this fraction of strongest peak clear_mz_range = 0.0 0.0 # for iTRAQ/TMT type data; will clear out all peaks in the specified m/z range

additional modifications

add_Cterm_peptide = 0.000000 add_Nterm_peptide = 0.000000 add_Cterm_protein = 0.000000 add_Nterm_protein = 0.000000

add_G_glycine = 0.000000 add_A_alanine = 0.000000 add_S_serine = 0.000000 add_P_proline = 0.000000 add_V_valine = 0.000000 add_T_threonine = 0.000000 add_C_cysteine = 57.021464 add_L_leucine = 0.000000 add_I_isoleucine = 0.000000 add_N_asparagine = 0.000000 add_D_aspartic_acid = 0.000000 add_Q_glutamine = 0.000000 add_K_lysine = 0.000000 add_E_glutamic_acid = 0.000000 add_M_methionine = 0.000000 add_H_histidine = 0.000000 add_F_phenylalanine = 0.000000 add_R_arginine = 0.000000 add_Y_tyrosine = 0.000000 add_W_tryptophan = 0.000000 add_B_user_amino_acid = 0.000000 add_J_user_amino_acid = 0.000000 add_O_user_amino_acid = 0.000000 add_U_user_amino_acid = 0.000000 add_X_user_amino_acid = 0.000000 add_Z_user_amino_acid = 0.000000

_ **

Run log output

** copy text from the text console in the gui here **

System info: System OS: Windows 7, Architecture: AMD64 Java Info: 1.8.0_211, Java HotSpot(TM) 64-Bit Server VM, Oracle Corporation

Version info: FragPipe version 9.4 MSFragger version 20190530 Philosopher version v1.0.1 (build 1558718856)

LCMS files: Experiment/Group:

12 commands to execute: Workspace [Work dir: C:\Users\Suzanne Ackloo\Desktop\Fragger output] C:\Users\Suzanne Ackloo\MSFragger-20190530\philosopher.exe workspace --clean Workspace [Work dir: C:\Users\Suzanne Ackloo\Desktop\Fragger output] C:\Users\Suzanne Ackloo\MSFragger-20190530\philosopher.exe workspace --init MsFragger [Work dir: C:\Users\Suzanne Ackloo\Desktop\Fragger output] java -jar -Dfile.encoding=UTF-8 -Xmx1G C:\Users\Suzanne Ackloo\MSFragger-20190530\MSFragger-20190530.jar C:\Users\Suzanne Ackloo\Desktop\Fragger output\fragger.params C:\Users\Suzanne Ackloo\Desktop\Digest test-60min_D1 soln_B6486.mzML C:\Users\Suzanne Ackloo\Desktop\Digest test-60min_D1C1 soln_A6483.mzML MsFragger java -cp C:\Users\Suzanne Ackloo\MSFragger-20190530\FragPipe.exe umich.msfragger.util.FileMove C:\Users\Suzanne Ackloo\Desktop\Digest test-60min_D1 soln_B6486.pepXML C:\Users\Suzanne Ackloo\Desktop\Fragger output\Digest test-60min_D1 soln_B6486.pepXML MsFragger java -cp C:\Users\Suzanne Ackloo\MSFragger-20190530\FragPipe.exe umich.msfragger.util.FileMove C:\Users\Suzanne Ackloo\Desktop\Digest test-60min_D1C1 soln_A6483.pepXML C:\Users\Suzanne Ackloo\Desktop\Fragger output\Digest test-60min_D1C1 soln_A6483.pepXML ReportDbAnnotate [Work dir: C:\Users\Suzanne Ackloo\Desktop\Fragger output] C:\Users\Suzanne Ackloo\MSFragger-20190530\philosopher.exe database --annotate C:\Users\Suzanne Ackloo\Documents\Fasta files\Fasta files\HUMANvalidated.fasta --prefix rev PeptideProphet [Work dir: C:\Users\Suzanne Ackloo\Desktop\Fragger output] C:\Users\Suzanne Ackloo\MSFragger-20190530\philosopher.exe peptideprophet --decoyprobs --ppm --accmass --nonparam --expectscore --decoy rev_ --database C:\Users\Suzanne Ackloo\Documents\Fasta files\Fasta files\HUMAN_validated.fasta Digest test-60min_D1 solnB6486.pepXML PeptideProphet [Work dir: C:\Users\Suzanne Ackloo\Desktop\Fragger output] C:\Users\Suzanne Ackloo\MSFragger-20190530\philosopher.exe peptideprophet --decoyprobs --ppm --accmass --nonparam --expectscore --decoy rev --database C:\Users\Suzanne Ackloo\Documents\Fasta files\Fasta files\HUMAN_validated.fasta Digest test-60min_D1C1 soln_A6483.pepXML ProteinProphet [Work dir: C:\Users\Suzanne Ackloo\Desktop\Fragger output] C:\Users\Suzanne Ackloo\MSFragger-20190530\philosopher.exe proteinprophet --maxppmdiff 2000000 C:\Users\Suzanne Ackloo\Desktop\Fragger output\interact-Digest test-60min_D1 soln_B6486.pep.xml C:\Users\Suzanne Ackloo\Desktop\Fragger output\interact-Digest test-60min_D1C1 solnA6483.pep.xml ReportFilter [Work dir: C:\Users\Suzanne Ackloo\Desktop\Fragger output] C:\Users\Suzanne Ackloo\MSFragger-20190530\philosopher.exe filter --sequential --razor --mapmods --prot 0.01 --tag rev --pepxml C:\Users\Suzanne Ackloo\Desktop\Fragger output --protxml C:\Users\Suzanne Ackloo\Desktop\Fragger output\interact.prot.xml ReportReport [Work dir: C:\Users\Suzanne Ackloo\Desktop\Fragger output] C:\Users\Suzanne Ackloo\MSFragger-20190530\philosopher.exe report Workspace [Work dir: C:\Users\Suzanne Ackloo\Desktop\Fragger output] C:\Users\Suzanne Ackloo\MSFragger-20190530\philosopher.exe workspace --clean


Workspace [Work dir: C:\Users\Suzanne Ackloo\Desktop\Fragger output]
C:\Users\Suzanne Ackloo\MSFragger-20190530\philosopher.exe workspace --clean
INFO[14:08:01] Executing Workspace v1.0.1                   
INFO[14:08:03] Removing workspace                           
INFO[14:08:03] Done                                         
Process 'Workspace' finished, exit code: 0

Workspace [Work dir: C:\Users\Suzanne Ackloo\Desktop\Fragger output]
C:\Users\Suzanne Ackloo\MSFragger-20190530\philosopher.exe workspace --init
INFO[14:08:03] Executing Workspace v1.0.1                   
INFO[14:08:04] Creating workspace                           
INFO[14:08:04] Done                                         
Process 'Workspace' finished, exit code: 0

MsFragger [Work dir: C:\Users\Suzanne Ackloo\Desktop\Fragger output]
java -jar -Dfile.encoding=UTF-8 -Xmx1G C:\Users\Suzanne Ackloo\MSFragger-20190530\MSFragger-20190530.jar C:\Users\Suzanne Ackloo\Desktop\Fragger output\fragger.params C:\Users\Suzanne Ackloo\Desktop\Digest test-60min_D1 soln_B6486.mzML C:\Users\Suzanne Ackloo\Desktop\Digest test-60min_D1C1 soln_A6483.mzML
MSFragger version MSFragger-20190530
MSFTBX version 1.9.10
(c) University of Michigan

System OS: Windows 7, Architecture: AMD64
Java Info: 1.8.0_211, Java HotSpot(TM) 64-Bit Server VM, Oracle Corporation
JVM started with 910 MB memory

***********************************FIRST SEARCH************************************
Parameters used in this search:
num_threads = 4
database_name = C:\Users\Suzanne Ackloo\Documents\Fasta files\Fasta files\HUMAN_validated.fasta
decoy_prefix = rev_
precursor_mass_lower = -30.0
precursor_mass_upper = 30.0
precursor_mass_units = 1
precursor_true_tolerance = 30.0
precursor_true_units = 1
fragment_mass_tolerance = 100.0
fragment_mass_units = 1
calibrate_mass = 2
evaluate_mass_calibration = false
isotope_error = 0/1
mass_offsets = 0
precursor_mass_mode = SELECTED
localize_delta_mass = false
delta_mass_exclude_ranges = (-1.5,3.5)
fragment_ion_series = b,y
search_enzyme_name = Trypsin
search_enzyme_cutafter = KR
search_enzyme_butnotafter = P
num_enzyme_termini = 2
allowed_missed_cleavage = 1
clip_nTerm_M = false
allow_multiple_variable_mods_on_residue = false
max_variable_mods_per_mod = 3
max_variable_mods_combinations = 5000
output_file_extension = pepXML
output_format = pepXML
output_report_topN = 1
output_max_expect = 50.0
report_alternative_proteins = false
override_charge = false
precursor_charge_low = 2
precursor_charge_high = 4
digest_min_length = 7
digest_max_length = 50
digest_mass_range_low = 200.0
digest_mass_range_high = 5000.0
max_fragment_charge = 3
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 = 100
minIonsScoring = 3
min_matched_fragments = 4
minimum_ratio = 0.01
clear_mz_range_low = 0.0
clear_mz_range_high = 0.0
variable_mod_01 = 15.99490 M
variable_mod_02 = 42.01060 [^
variable_mod_03 = 79.96633 STY
Number of unique peptides
    of length 7: 123460
    of length 8: 114318
    of length 9: 108692
    of length 10: 100719
    of length 11: 93074
    of length 12: 86081
    of length 13: 81536
    of length 14: 73427
    of length 15: 69251
    of length 16: 63252
    of length 17: 57909
    of length 18: 54561
    of length 19: 50371
    of length 20: 46341
    of length 21: 43130
    of length 22: 39597
    of length 23: 37183
    of length 24: 34815
    of length 25: 32037
    of length 26: 29648
    of length 27: 28108
    of length 28: 26394
    of length 29: 23979
    of length 30: 22392
    of length 31: 20556
    of length 32: 19144
    of length 33: 17696
    of length 34: 16498
    of length 35: 15033
    of length 36: 14322
    of length 37: 12736
    of length 38: 11904
    of length 39: 10752
    of length 40: 10226
    of length 41: 9543
    of length 42: 8614
    of length 43: 7482
    of length 44: 6104
    of length 45: 4843
    of length 46: 3293
    of length 47: 2120
    of length 48: 1260
    of length 49: 716
    of length 50: 395
In total 1633512 peptides.
Exception in thread "main" java.lang.reflect.InvocationTargetException
    at sun.reflect.NativeMethodAccessorImpl.invoke0(Native Method)
    at sun.reflect.NativeMethodAccessorImpl.invoke(Unknown Source)
    at sun.reflect.DelegatingMethodAccessorImpl.invoke(Unknown Source)
    at java.lang.reflect.Method.invoke(Unknown Source)
    at com.simontuffs.onejar.Boot.run(Boot.java:340)
    at com.simontuffs.onejar.Boot.main(Boot.java:166)
Caused by: java.util.concurrent.ExecutionException: java.lang.OutOfMemoryError: Java heap space
    at java.util.concurrent.FutureTask.report(Unknown Source)
    at java.util.concurrent.FutureTask.get(Unknown Source)
    at edu.umich.andykong.msfragger.a.a(Unknown Source)
    at edu.umich.andykong.msfragger.K.a(Unknown Source)
    at edu.umich.andykong.msfragger.q.a(Unknown Source)
    at edu.umich.andykong.msfragger.B.<init>(Unknown Source)
    at edu.umich.andykong.msfragger.MSFragger.a(Unknown Source)
    at edu.umich.andykong.msfragger.MSFragger.main(Unknown Source)
    ... 6 more
Caused by: java.lang.OutOfMemoryError: Java heap space
    at edu.umich.andykong.msfragger.b.a(Unknown Source)
    at edu.umich.andykong.msfragger.a.a(Unknown Source)
    at edu.umich.andykong.msfragger.a$$Lambda$22/1161082381.call(Unknown Source)
    at java.util.concurrent.FutureTask.run(Unknown Source)
    at java.util.concurrent.ThreadPoolExecutor.runWorker(Unknown Source)
    at java.util.concurrent.ThreadPoolExecutor$Worker.run(Unknown Source)
    at java.lang.Thread.run(Unknown Source)
Process 'MsFragger' finished, exit code: 1

Process returned non-zero exit code, stopping
MsFragger
java -cp C:\Users\Suzanne Ackloo\MSFragger-20190530\FragPipe.exe umich.msfragger.util.FileMove C:\Users\Suzanne Ackloo\Desktop\Digest test-60min_D1 soln_B6486.pepXML C:\Users\Suzanne Ackloo\Desktop\Fragger output\Digest test-60min_D1 soln_B6486.pepXML

Stopping 10/13 processes (3 already finished) Processing interrupted, stopping MsFragger

fcyu commented 5 years ago

Hi,

From the command 

java -jar -Dfile.encoding=UTF-8 -Xmx1G C:\Users\Suzanne Ackloo\MSFragger-20190530\MSFragger-20190530.jar C:\Users\Suzanne Ackloo\Desktop\Fragger output\fragger.params C:\Users\Suzanne Ackloo\Desktop\Digest test-60min_D1 soln_B6486.mzML C:\Users\Suzanne Ackloo\Desktop\Digest test-60min_D1C1 soln_A6483.mzML

you only provided 1GB to java, which was far from enough. I suggest you specifying as much as your PC have to RAM (GB) in the MSFragger tab of FragPipe.

Bests,

Fengchao

guoci commented 5 years ago

Please see https://github.com/Nesvilab/FragPipe/issues/113#issuecomment-487608742