Hi Sarah, Can you send me your protein list and an excerpt from your protein database? I still suspect it's some sort of identifier mismatch issue, especially given that your peptides/transitions are lost at the step of creating the protein-restricted library.
Thanks! Carly On Oct 7, 2:09 pm, Sarah <[email protected]> wrote: > Hi Carly, > > thanks for your ideas. Unfortunately, still no transitions on my > list....I did make sure the "protein" box was checked and also created > a new list with IPI identifiers (the IPI identifier in the format > IPI00123456 is sufficient, right?) to double check the format. This is > the log for this search: > > # Commands for session MV240KSZQ on Wed Oct 7 16:44:07 2009 > # BEGIN COMMAND BLOCK > ###### BEGIN Command Execution ###### > [Wed Oct 7 16:44:08 2009] EXECUTING: run_marimba.pl -t10 -s3 -k -m300 > -M1200 -R c:/Inetpub/wwwroot/ISB/data/IPIwhole.txt -X C,M -T Q -i y -z > 2,3 -Z 1,2 c:/Inetpub/wwwroot/ISB/data/MaRiMba/ > NIST_human_IT_v3.0_2009_02_04_7AA.splib c:/Inetpub/wwwroot/ISB/data/ > ipi.HUMAN.v3.64.fasta c:/Inetpub/wwwroot/ISB/data/MRMlist.txt > OUTPUT: > Run MaRiMba started at: Wed Oct 7 16:44:08 2009 > ...Refreshing library against database > ...Filtering out non-proteotypic and unmapped peptides in a tryptic > context only > SpectraST started at Wed Oct 07 16:44:08 2009. > Creating library from "c:/Inetpub/wwwroot/ISB/data/MaRiMba/ > NIST_human_IT_v3.0_2009_02_04_7AA.splib" > REFRESHING protein mappings...DONE! > Importing peptide ions... > 500...1000...1500...2000...2500...3000...3500...4000...4500...5000...5500...6000...6500...7000...7500...8000...8500...9000...9500...10000...10500...11000...11500...12000...12500...13000...13500...14000...14500...15000...15500...16000...16500...17000...17500...18000...18500...19000...19500...20000...20500...21000...21500...22000...22500...23000...23500...24000...24500...25000...25500...26000...26500...27000...27500...28000...28500...29000...29500...30000...30500...31000...31500...32000...32500...33000...33500...34000...34500...35000...35500...36000...36500...37000...37500...38000...38500...39000...39500...40000...40500...41000...41500...42000...42500...43000...43500...44000...44500...45000...45500...46000...46500...47000...47500...48000...48500...49000...49500...50000...50500...51000...51500...52000...52500...53000...53500...54000...54500...55000...55500...56000...56500...57000...57500...58000...58500...59000...59500...60000...60500...61000...61500...62000...62500...63000...63500...64000...64500...65000...65500...66000...66500...67000...67500...68000...68500...69000...69500...70000...70500...71000...71500...72000...72500...73000...73500...74000...74500...75000...75500...76000...76500...77000...77500...78000...78500...79000...79500...80000...80500...81000...81500...82000...82500...83000...83500...84000...84500...85000...85500...86000...86500...87000...87500...88000...88500...89000...89500...90000...90500...91000...91500...92000...92500...93000...93500...94000...94500...95000...95500...96000...96500...97000...97500...98000...98500...99000...99500...100000...100500...101000...101500...102000...102500...103000...103500...104000...104500...105000...105500...106000...106500...107000...107500...108000...108500...109000...109500...110000...110500...111000...111500...112000...112500...113000...113500...114000...114500...115000...115500...116000...116500...117000...117500...118000...118500...119000...119500...120000...120500...121000...121500...122000...122500...123000...123500...124000...124500...125000...125500...126000...126500...127000...127500...128000...128500...129000...129500...130000...130500...131000...131500...132000...132500...133000...133500...134000...134500...135000...135500...136000...136500...137000...137500...138000...138500...139000...139500...140000...140500...141000...141500...142000...142500...143000...143500...144000...144500...145000...145500...146000...146500...147000...147500...148000...148500...149000...149500...150000...150500...151000...151500...152000...152500...153000...153500...154000...154500...155000...155500...156000...156500...157000...157500...158000...158500...159000...159500...160000...160500...161000...161500...162000...162500...163000...163500...164000...164500...165000...165500...166000...166500...167000...167500...168000...168500...169000...169500...170000...170500...171000...171500...172000...172500...173000...173500...174000...174500...175000...175500...176000...176500...177000...177500...178000...178500...179000...179500...180000...180500...181000...181500...182000...182500...183000...183500...184000...184500...185000...185500...186000...186500...187000...187500...188000...188500...189000...189500...190000...190500...191000...191500...192000...192500...193000...193500...194000...194500...195000...195500...196000...196500...197000...197500...198000...198500...199000...199500...200000...200500...201000...201500...202000...202500...203000...203500...204000...204500...205000...205500...206000...206500...207000...207500...208000...208500...209000...209500...210000...210500...211000...211500...212000...212500...213000...213500...214000...214500...215000...215500...216000...216500...217000...217500...218000...218500...219000...219500...220000...220500...221000...221500...222000...222500...223000...223500...224000...224500...225000...225500...226000...226500...227000...227500...228000...228500...229000...229500...230000...230500...231000...231500...232000...232500...233000...233500...234000...234500...235000...235500...236000...236500...237000...237500...238000...238500...239000...239500...240000...240500...241000...241500...242000...242500...243000...243500...244000...244500...245000...245500...246000...246500...247000...247500...248000...248500...249000...249500...250000...250500...251000...251500...252000...252500...253000...253500...254000...254500...255000...255500...256000...256500...257000...257500...258000...258500...259000...259500...260000...260500...DONE! > > Library file (BINARY) "tmp_refreshed.splib" created. > Library file (TEXT) "tmp_refreshed.sptxt" created. > M/Z Index file "tmp_refreshed.spidx" created. > Peptide Index file "tmp_refreshed.pepidx" created. > > Total number of spectra in library: 63094 > Total number of distinct peptide ions in library: 63094 > Total number of distinct stripped peptides in library: 37177 > CHARGE +1: 4595 ; +2: 34631 ; +3: 20407 > TERMINI Tryptic: 63094 ; Semi-tryptic: 0 ; Non-tryptic: 0 > METHIONINE MOD Oxidized: 8160 > CYSTEINE MOD CAM: 9231 ; Cleavable-ICAT: 1181 ; Uncleavable-ICAT: > 1453 > PROBABILITY >0.9999: 47423 ; 0.999-0.9999: 9010 ; 0.99-0.999: > 4724 ; 0.9-0.99: 1937 <0.9: 0 > NREPS 20+: 11473 ; 10-19: 8857 ; 4-9: 18596 ; 2-3: 24168 ; > 1: 0 > > Total Run Time = 938 seconds. > SpectraST finished at Wed Oct 07 16:59:46 2009 without error. > ...Restricting input library to proteins on list > SpectraST started at Wed Oct 07 16:59:46 2009. > Creating library from "c://tmp_refreshed.splib" > Importing peptide ions... > 500...1000...1500...2000...2500...3000...3500...4000...4500...5000...5500...6000...6500...7000...7500...8000...8500...9000...9500...10000...10500...11000...11500...12000...12500...13000...13500...14000...14500...15000...15500...16000...16500...17000...17500...18000...18500...19000...19500...20000...20500...21000...21500...22000...22500...23000...23500...24000...24500...25000...25500...26000...26500...27000...27500...28000...28500...29000...29500...30000...30500...31000...31500...32000...32500...33000...33500...34000...34500...35000...35500...36000...36500...37000...37500...38000...38500...39000...39500...40000...40500...41000...41500...42000...42500...43000...43500...44000...44500...45000...45500...46000...46500...47000...47500...48000...48500...49000...49500...50000...50500...51000...51500...52000...52500...53000...53500...54000...54500...55000...55500...56000...56500...57000...57500...58000...58500...59000...59500...60000...60500...61000...61500...62000...62500...63000...DONE! > > Library file (BINARY) "tmp_restricted.splib" created. > Library file (TEXT) "tmp_restricted.sptxt" created. > M/Z Index file "tmp_restricted.spidx" created. > Peptide Index file "tmp_restricted.pepidx" created. > > Total number of spectra in library: 0 > Total number of distinct peptide ions in library: 0 > Total number of distinct stripped peptides in library: 0 > CHARGE +1: 0 ; +2: 0 ; +3: 0 > TERMINI Tryptic: 0 ; Semi-tryptic: 0 ; Non-tryptic: 0 > METHIONINE MOD Oxidized: 0 > CYSTEINE MOD CAM: 0 ; Cleavable-ICAT: 0 ; Uncleavable-ICAT: 0 > PROBABILITY >0.9999: 0 ; 0.999-0.9999: 0 ; 0.99-0.999: 0 ; > 0.9-0.99: 0 <0.9: 0 > NREPS 20+: 0 ; 10-19: 0 ; 4-9: 0 ; 2-3: 0 ; 1: 0 > > Total Run Time = 110 seconds. > SpectraST finished at Wed Oct 07 17:01:36 2009 without error. > ...Creating consensus library > SpectraST started at Wed Oct 07 17:01:36 2009. > Creating CONSENSUS library from "c://tmp_restricted.splib" > Importing peptide ions...DONE! > > Library file (BINARY) "tmp_consensus.splib" created. > Library file (TEXT) "tmp_consensus.sptxt" created. > M/Z Index file "tmp_consensus.spidx" created. > Peptide Index file "tmp_consensus.pepidx" created. > > Total number of spectra in library: 0 > Total number of distinct peptide ions in library: 0 > Total number of distinct stripped peptides in library: 0 > CHARGE +1: 0 ; +2: 0 ; +3: 0 > TERMINI Tryptic: 0 ; Semi-tryptic: 0 ; Non-tryptic: 0 > METHIONINE MOD Oxidized: 0 > CYSTEINE MOD CAM: 0 ; Cleavable-ICAT: 0 ; Uncleavable-ICAT: 0 > PROBABILITY >0.9999: 0 ; 0.999-0.9999: 0 ; 0.99-0.999: 0 ; > 0.9-0.99: 0 <0.9: 0 > NREPS 20+: 0 ; 10-19: 0 ; 4-9: 0 ; 2-3: 0 ; 1: 0 > > Total Run Time = 0 seconds. > SpectraST finished at Wed Oct 07 17:01:36 2009 without error. > ...Applying quality filters > SpectraST started at Wed Oct 07 17:01:36 2009. > Applying QUALITY FILTER to "tmp_consensus.splib". > Importing peptide ions...DONE! > > Library file (BINARY) "tmp_consensus_Q2.splib" created. > Library file (TEXT) "tmp_consensus_Q2.sptxt" created. > M/Z Index file "tmp_consensus_Q2.spidx" created. > Peptide Index file "tmp_consensus_Q2.pepidx" created. > > Total number of spectra in library: 0 > Total number of distinct peptide ions in library: 0 > Total number of distinct stripped peptides in library: 0 > CHARGE +1: 0 ; +2: 0 ; +3: 0 > TERMINI Tryptic: 0 ; Semi-tryptic: 0 ; Non-tryptic: 0 > METHIONINE MOD Oxidized: 0 > CYSTEINE MOD CAM: 0 ; Cleavable-ICAT: 0 ; Uncleavable-ICAT: 0 > PROBABILITY >0.9999: 0 ; 0.999-0.9999: 0 ; 0.99-0.999: 0 ; > 0.9-0.99: 0 <0.9: 0 > NREPS 20+: 0 ; 10-19: 0 ; 4-9: 0 ; 2-3: 0 ; 1: 0 > > Total Run Time = 0 seconds. > SpectraST finished at Wed Oct 07 17:01:36 2009 without error. > ...Performing initial peptide filtering > SpectraST started at Wed Oct 07 17:01:36 2009. > Creating library from "c://tmp_consensus_Q2.splib" > Importing peptide ions...DONE! > > Library file (BINARY) "tmp_filtered.splib" created. > Library file (TEXT) "tmp_filtered.sptxt" created. > M/Z Index file "tmp_filtered.spidx" created. > Peptide Index file "tmp_filtered.pepidx" created. > > Total number of spectra in library: 0 > Total number of distinct peptide ions in library: 0 > Total number of distinct stripped peptides in library: 0 > CHARGE +1: 0 ; +2: 0 ; +3: 0 > TERMINI Tryptic: 0 ; Semi-tryptic: 0 ; Non-tryptic: 0 > METHIONINE MOD Oxidized: 0 > CYSTEINE MOD CAM: 0 ; Cleavable-ICAT: 0 ; Uncleavable-ICAT: 0 > PROBABILITY >0.9999: 0 ; 0.999-0.9999: 0 ; 0.99-0.999: 0 ; > 0.9-0.99: 0 <0.9: 0 > NREPS 20+: 0 ; 10-19: 0 ; 4-9: 0 ; 2-3: 0 ; 1: 0 > > Total Run Time = 0 seconds. > SpectraST finished at Wed Oct 07 17:01:36 2009 without error. > ...Creating preliminary MRM list > SpectraST started at Wed Oct 07 17:01:36 2009. > Creating library from "c://tmp_filtered.splib" > Importing peptide ions...DONE! > > Library file (BINARY) "tmp_MRM.splib" created. > Library file (TEXT) "tmp_MRM.sptxt" created. > M/Z Index file "tmp_MRM.spidx" created. > Peptide Index file "tmp_MRM.pepidx" created. > MRM Table file "tmp_MRM.mrm" created. > > Total number of spectra in library: 0 > Total number of distinct peptide ions in library: 0 > Total number of distinct stripped peptides in library: 0 > CHARGE +1: 0 ; +2: 0 ; +3: 0 > TERMINI Tryptic: 0 ; Semi-tryptic: 0 ; Non-tryptic: 0 > METHIONINE MOD Oxidized: 0 > CYSTEINE MOD CAM: 0 ; Cleavable-ICAT: 0 ; Uncleavable-ICAT: 0 > PROBABILITY >0.9999: 0 ; 0.999-0.9999: 0 ; 0.99-0.999: 0 ; > 0.9-0.99: 0 <0.9: 0 > NREPS 20+: 0 ; 10-19: 0 ; 4-9: 0 ; 2-3: 0 ; 1: 0 > > Total Run Time = 0 seconds. > SpectraST finished at Wed Oct 07 17:01:36 2009 without error. > ...Applying user-defined fiters > ...Sorting results > ...Restricting to minimum number of transitions per peptide > ...Deleting temporary files > Job finished at: Wed Oct 7 17:01:38 2009 > > END OUTPUT > RETURN CODE:0 > ###### End Command Execution ###### > # All finished at Wed Oct 7 17:01:38 2009 > # END COMMAND BLOCK > > Does that tell you anything? > > Thank you! > Sarah > > On Oct 7, 3:17 pm, Carly <[email protected]> wrote:> Hi Sarah, > > > Try double-checking two things: > > > 1. In the Filtering Options section, ensure that "Proteins" is > > selected where it says, "Please specify whether the restriction file > > listed in step 3 above contains Proteins or Peptides." > > > 2. Make sure your protein restriction list contains only a single > > column of IPI identifiers, e.g.: > > > IPI00123456 > > IPI00123457 > > IPI00123458 > > IPI00123956 > > IPI00133456 > > > If neither of these steps does the trick, please post the complete > > thread of MaRiMba output (i.e. including the 102 lines that it skips > > in the abridged version), so I can have a better idea of where the > > problem is occurring. > > > Hope this helps! > > > Carly > > Martin Group > > Institute for Systems Biology > > > On Oct 7, 8:40 am, Sarah <[email protected]> wrote: > > > > Hey all, > > > > I have to say I am not too tech-savvy but think I took the right steps > > > in trying to create a list of MRM transitions with the MaRiMba tool. > > > > I have installed TPP v4.1 MaRiMba rev1 (on a windows operating system) > > > and it does create me a MRM list when I feed in the spectral library > > > and database with the mix of 18 proteins that were included as files > > > within the software package. > > > It also gives out MRM transitions when I put in as spectral library > > > NIST_human_IT_v3.0_2009_02_04_7AA.splib and as database the human IPI > > > database (ipi.HUMAN.v3.64.fasta) without restricting to specific > > > proteins. > > > But then when I specify a protein restriction list, I always end up > > > with 0 transitions. I do use the same format as the input ipi and I > > > even copied the whole ipi file from the database into a text file > > > (.txt) to make sure I have the exact same format as the database. But > > > still 0 mrms... > > > > This is the output I got with the above input: > > > > run_marimba.pl -t10 -s3 -k -m300 -M1200 -R c:/Inetpub/wwwroot/ISB/data/ > > > complete.txt -X C,M -T Q -i y -z 2,3 -Z 1,2 c:/Inetpub/wwwroot/ISB/ > > > data/MaRiMba/NIST_human_IT_v3.0_2009_02_04_7AA.splib c:/Inetpub/ > > > wwwroot/ISB/data/ipi.HUMAN.v3.64.fasta c:/Inetpub/wwwroot/ISB/data/ > > > MRMlist.txt > > > ________________________________________ > > > Run MaRiMba started at: Wed Oct 7 09:31:42 2009 > > > ...Refreshing library against database > > > ...Filtering out non-proteotypic and unmapped peptides in a tryptic > > > context only > > > SpectraST started at Wed Oct 07 09:31:42 2009. > > > Creating library from "c:/Inetpub/wwwroot/ISB/data/MaRiMba/ > > > NIST_human_IT_v3.0_2009_02_04_7AA.splib" > > > REFRESHING protein mappings...DONE! > > > Importing peptide ions... > > > 500...1000...1500...2000...2500...3000...3500...4000...4500...5000...5500...6000...6500...7000...7500...8000...8500...9000...9500...10000...10500...11000...11500...12000...12500...13000...13500...14000...14500...15000...15500...16000...16500...17000...17500...18000...18500...19000...19500...20000...20500...21000...21500...22000...22500...23000...23500...24000...24500...25000...25500...26000...26500...27000...27500...28000...28500...29000...29500...30000...30500...31000...31500...32000...32500...33000...33500...34000...34500...35000...35500...36000...36500...37000...37500...38000...38500...39000...39500...40000...40500...41000...41500...42000...42500...43000...43500...44000...44500...45000...45500...46000...46500...47000...47500...48000...48500...49000...49500...50000...50500...51000...51500...52000...52500...53000...53500...54000...54500...55000...55500...56000...56500...57000...57500...58000...58500...59000...59500...60000...60500...61000...61500...62000...62500...63000...63500...64000...64500...65000...65500...66000...66500...67000...67500...68000...68500...69000...69500...70000...70500...71000...71500...72000...72500...73000...73500...74000...74500...75000...75500...76000...76500...77000...77500...78000...78500...79000...79500...80000...80500...81000...81500...82000...82500...83000...83500...84000...84500...85000...85500...86000...86500...87000...87500...88000...88500...89000...89500...90000...90500...91000...91500...92000...92500...93000...93500...94000...94500...95000...95500...96000...96500...97000...97500...98000...98500...99000...99500...100000...100500...101000...101500...102000...102500...103000...103500...104000...104500...105000...105500...106000...106500...107000...107500...108000...108500...109000...109500...110000...110500...111000...111500...112000...112500...113000...113500...114000...114500...115000...115500...116000...116500...117000...117500...118000...118500...119000...119500...120000...120500...121000...121500...122000...122500...123000...123500...124000...124500...125000...125500...126000...126500...127000...127500...128000...128500...129000...129500...130000...130500...131000...131500...132000...132500...133000...133500...134000...134500...135000...135500...136000...136500...137000...137500...138000...138500...139000...139500...140000...140500...141000...141500...142000...142500...143000...143500...144000...144500...145000...145500...146000...146500...147000...147500...148000...148500...149000...149500...150000...150500...151000...151500...152000...152500...153000...153500...154000...154500...155000...155500...156000...156500...157000...157500...158000...158500...159000...159500...160000...160500...161000...161500...162000...162500...163000...163500...164000...164500...165000...165500...166000...166500...167000...167500...168000...168500...169000...169500...170000...170500...171000...171500...172000...172500...173000...173500...174000...174500...175000...175500...176000...176500...177000...177500...178000...178500...179000...179500...180000...180500...181000...181500...182000...182500...183000...183500...184000...184500...185000...185500...186000...186500...187000...187500...188000...188500...189000...189500...190000...190500...191000...191500...192000...192500...193000...193500...194000...194500...195000...195500...196000...196500...197000...197500...198000...198500...199000...199500...200000...200500...201000...201500...202000...202500...203000...203500...204000...204500...205000...205500...206000...206500...207000...207500...208000...208500...209000...209500...210000...210500...211000...211500...212000...212500...213000...213500...214000...214500...215000...215500...216000...216500...217000...217500...218000...218500...219000...219500...220000...220500...221000...221500...222000...222500...223000...223500...224000...224500...225000...225500...226000...226500...227000...227500...228000...228500...229000...229500...230000...230500...231000...231500...232000...232500...233000...233500...234000...234500...235000...235500...236000...236500...237000...237500...238000...238500...239000...239500...240000...240500...241000...241500...242000...242500...243000...243500...244000...244500...245000...245500...246000...246500...247000...247500...248000...248500...249000...249500...250000...250500...251000...251500...252000...252500...253000...253500...254000...254500...255000...255500...256000...256500...257000...257500...258000...258500...259000...259500...260000...260500...DONE! > > > > Library file (BINARY) "tmp_refreshed.splib" created. > > > Library file (TEXT) "tmp_refreshed.sptxt" created. > > > M/Z Index file "tmp_refreshed.spidx" created. > > > Peptide Index file "tmp_refreshed.pepidx" created. > > > > Total number of spectra in library: 63094 > > > Total number of distinct peptide ions in library: 63094 > > > Total number of distinct stripped peptides in library: 37177 > > > CHARGE +1: 4595 ; +2: 34631 ; +3: 20407 > > > TERMINI Tryptic: 63094 ; Semi-tryptic: 0 ; Non-tryptic: 0 > > > METHIONINE MOD Oxidized: 8160 > > > CYSTEINE MOD CAM: 9231 ; Cleavable-ICAT: 1181 ; Uncleavable-ICAT: > > > 1453 > > > > ________________________________________ > > > ...skipping 102 lines of output... > > > ________________________________________ > > > MRM Table file "tmp_MRM.mrm" created. > > > > Total number of spectra in library: 0 > > > Total number of distinct peptide ions in library: 0 > > > Total number of distinct stripped peptides in library: 0 > > > CHARGE +1: 0 ; +2: 0 ; +3: 0 > > > TERMINI Tryptic: 0 ; Semi-tryptic: 0 ; Non-tryptic: 0 > > > METHIONINE MOD Oxidized: 0 > > > CYSTEINE MOD CAM: 0 ; Cleavable-ICAT: 0 ; Uncleavable-ICAT: 0 > > > PROBABILITY >0.9999: 0 ; 0.999-0.9999: 0 ; 0.99-0.999: 0 ; > > > 0.9-0.99: 0 <0.9: 0 > > > NREPS 20+: 0 ; 10-19: 0 ; 4-9: 0 ; 2-3: 0 ; 1: 0 > > > > Total Run Time = 0 seconds. > > > SpectraST finished at Wed Oct 07 09:48:09 2009 without error. > > > ...Applying user-defined fiters > > > ...Sorting results > > > ...Restricting to minimum number of transitions per peptide > > > ...Deleting temporary files > > > Job finished at: Wed Oct 7 09:48:12 2009 > > > > Command Successful > > > > I don't know if that helps.. > > > > I would really appreciate any ideas!! > > > > Thanks, Sarah --~--~---------~--~----~------------~-------~--~----~ You received this message because you are subscribed to the Google Groups "spctools-discuss" group. To post to this group, send email to [email protected] To unsubscribe from this group, send email to [email protected] For more options, visit this group at http://groups.google.com/group/spctools-discuss?hl=en -~----------~----~----~----~------~----~------~--~---
