Hi Brian, Glad that you like the new (still beta) protXML viewer. Just as a quick update, the version shipped with 4.7.1 has links to the spectra (via the peptide sequence link), as well as to all of the proteins that a given peptide sequence is shared by (via the weight link). Quantitation values and links for XPRESS and ASAPRatio are also shown; Libra needs a little work still. You can also export a basic tab-delimited file suitable for Excel.
We plan on making this viewer the default in our next release; do let us know of any bugs or desired features in the meantime! Cheers, --Luis On Wed, Mar 5, 2014 at 3:51 PM, Brian Hampton <[email protected]> wrote: > Hi David, > > You are right, searching all of Uniprot + decoys does take a while, about > 6 hours for a relatively small data set. > Unfortunately, in my case, there were only a few sequences in there from > the species being studied. Apparently only reviewed sequences are in the > "full" uniprot database, or at least in the version I downloaded. > > Anyway, I guessed correctly on the "contaminating" species in the sample. > I concatenated that database with the database for the species under study > and viola the pep.xml files run nicely through the TPP with many hundreds > matching to proteins from the offending species database. In the end, only > two proteins matched to the database in the species under study and 650 > matched to the offending species. So no wonder TPP couldn't give me any > validated results. > > I did this using the latest version of TPP (4.7) as you suggested. I like > the beta version of the viewer for the prot.xml files. Though I haven't > found a link from that view to see the ms/ms spectra. Also the iProphet > results are now fully viewable (on linux). Before only the first line > would be displayed. > > Thanks a bunch for your help! > > Cheers! > Brian > > > > On Sat, Mar 1, 2014 at 12:01 PM, David Shteynberg < > [email protected]> wrote: > >> Hi Brian, >> >> I have recently been confirming that incomplete databases could pose a >> big problem to properly modeling the data. One solution is to search >> against all of uniprot plus decoys. The good part about this is you don't >> have to include contaminants as they should already be represented ;-). >> The bad part is the searches will take longer. >> >> -David >> >> >> >> >> >> >> On Sat, Mar 1, 2014 at 6:44 AM, Brian Hampton <[email protected]>wrote: >> >>> Thanks David, I'll try your suggestions. Also, I'm thinking there might >>> be proteins from another species in the sample that are not represented in >>> the database I'm searching. >>> >>> Thanks, >>> Brian >>> >>> On Feb 28, 2014, at 6:18 PM, David Shteynberg < >>> [email protected]> wrote: >>> >>> Hi Brian, >>> >>> This message is quite telling: 10751 Decoys, and 11603 Non-Decoys >>> >>> Assuming 50% of the wrong ones are Decoys there are less than 900 >>> correct matches in this dataset. >>> >>> >>> That said, I don't think using the default parameteric option is the >>> best choice here due to the extended tail of the negative distribution and >>> the relatively small number of correct IDs. I would suggest you try the >>> semi-parametric (non-parametric option), also run with minimum probability >>> 0 and perhaps set a CLEVEL of 1 with advanced option -c1 or -c2 (higher is >>> more conservative) to help keep the positive distribution of the model on >>> the shoulder. >>> >>> Finally, I would recommend you try the latest version 4.7.0, which is >>> due out today. >>> >>> Cheers, >>> -David >>> >>> >>> On Fri, Feb 28, 2014 at 3:09 PM, Brian Hampton <[email protected]>wrote: >>> >>>> I am trying to squeak additional mass accuracy out of an LTQ by >>>> collecting MS1 data with Enhanced Scan and in Profile mode so the data will >>>> have isotopic resolution. This had the expected result of shifting the >>>> frequency of peptides observed at a particular error down from ~0.75 (when >>>> collected using normal scan rate and centroid mode) to center the curve >>>> over an error of 0 (when collected using enhanced scan rate and profile >>>> mode). Nice. >>>> >>>> I processed the raw files with msconvert and used the --filter >>>> "peakPicking true 1-1" argument and run it through TPP v 4.6.2 with Tandem >>>> as the search engine. This has worked fine on samples until today when >>>> this latest data set (which is a pull down experiment), xinteract fails to >>>> produce a pep.xml file and results in a warning that says: >>>> >>>> WARNING: Mixture model quality test failed for charge (1+). >>>> WARNING: Mixture model quality test failed for charge (2+). >>>> WARNING: Mixture model quality test failed for charge (3+). >>>> >>>> The tandem.pep.xml file contains many high scoring peptides. And there >>>> are good +1,+2 & +3 spectral matches. >>>> >>>> I am wondering if there is an argument to msconvert that I am missing. >>>> Or maybe my approach to collecting the data is incompatible with TPP? Or >>>> could this be a problem with K-score vs native tandem scoring? I haven't >>>> tried another search using native Tandem scoring yet. >>>> >>>> Below is the output from TPP. >>>> >>>> Thanks in advance for any help. >>>> >>>> Cheers, >>>> Brian >>>> >>>> >>>> >>>> >>>> >>>> EXECUTING: cd /usr/local/tpp/data/projects/lindsey/xiap-ide12; >>>> /usr/local/tpp/bin/xinteract -Ninteract-XIAP-IDE12.pep.xml -p0.05 -l5 -Op >>>> -dDECOY 140226-XIAP-IDE12-5pct.tandem.pep.xml >>>> >>>> /usr/local/tpp/bin/xinteract (TPP v4.6 OCCUPY rev 2, Build 201302151642 >>>> (linux)) >>>> >>>> running: "/usr/local/tpp/bin/InteractParser >>>> 'interact-XIAP-IDE12.pep.xml' '140226-XIAP-IDE12-5pct.tandem.pep.xml' >>>> -L'5'" >>>> file 1: 140226-XIAP-IDE12-5pct.tandem.pep.xml >>>> SUCCESS: CORRECTED data file >>>> /usr/local/tpp/data/projects/lindsey/xiap-ide12/140226-XIAP-IDE12-5pct.mzML >>>> in msms_run_summary tag ... >>>> processed altogether 22365 results >>>> INFO: Results written to file: >>>> /usr/local/tpp/data/projects/lindsey/xiap-ide12/interact-XIAP-IDE12.pep.xml >>>> command completed in 6 sec >>>> >>>> running: "/usr/local/tpp/bin/DatabaseParser >>>> 'interact-XIAP-IDE12.pep.xml'" >>>> command completed in 1 sec >>>> >>>> running: "/usr/local/tpp/bin/RefreshParser >>>> 'interact-XIAP-IDE12.pep.xml' >>>> '/usr/local/tpp/data/dbase/ixodes-plus-crap_DECOY.fasta'" >>>> - Building Commentz-Walter keyword tree... - Searching the tree... >>>> - Linking duplicate entries... - Printing results... >>>> >>>> command completed in 6 sec >>>> >>>> running: "/usr/local/tpp/bin/PeptideProphetParser >>>> 'interact-XIAP-IDE12.pep.xml' MINPROB=0.05 DECOY=DECOY" >>>> Using Decoy Label "DECOY". >>>> (X! Tandem (k-score)) >>>> init with X! Tandem (k-score) trypsin >>>> MS Instrument info: Manufacturer: Thermo Scientific, Model: UNKNOWN, >>>> Ionization: nanoelectrospray, Analyzer: radial ejection linear ion trap, >>>> Detector: electron multiplier >>>> >>>> PeptideProphet (TPP v4.6 OCCUPY rev 2, Build 201302151642 (linux)) >>>> AKeller@ISB >>>> read in 2445 1+, 9959 2+, 9950 3+, 0 4+, 0 5+, 0 6+, and 0 7+ spectra. >>>> Initialising statistical models ... >>>> Found 10751 Decoys, and 11603 Non-Decoys >>>> Iterations: .........10.........20........ >>>> WARNING: Mixture model quality test failed for charge (1+). >>>> WARNING: Mixture model quality test failed for charge (2+). >>>> WARNING: Mixture model quality test failed for charge (3+). >>>> model complete after 29 iterations >>>> command completed in 4 sec >>>> >>>> running: "/usr/local/tpp/bin/ProphetModels.pl -i >>>> interact-XIAP-IDE12.pep.xml -d DECOY" >>>> Analyzing interact-XIAP-IDE12.pep.xml ... >>>> Parsing search results >>>> "/usr/local/tpp/data/projects/lindsey/xiap-ide12/140226-XIAP-IDE12-5pct (X! >>>> Tandem (k-score))"... >>>> => Total of 0 hits. >>>> => Total of 0 decoy hits. >>>> => Total of 0 excluded hits. >>>> Warning: empty y range [0:0], adjusting to [0:1] >>>> Warning: empty y range [0:0], adjusting to [0:1] >>>> >>>> plot "interact-XIAP-IDE12.pep_FVAL.tsv" using 1:17 title "Observed" >>>> with line lc -1 , "interact-XIAP-IDE12.pep_FVAL.tsv" using 1:18 title >>>> "Model Pos" with line lc 3 , "interact-XIAP-IDE12.pep_FVAL.tsv" using 1:19 >>>> title "Model Neg" with line lc 1 >>>> >>>> ^ >>>> "interact-XIAP-IDE12.pep_FVAL.gp", line 23: warning: Skipping data >>>> file with no valid points >>>> Warning: empty y range [0:0], adjusting to [0:1] >>>> >>>> plot "interact-XIAP-IDE12.pep_FVAL.tsv" using 1:20 title "Observed" >>>> with line lc -1 , "interact-XIAP-IDE12.pep_FVAL.tsv" using 1:21 title >>>> "Model Pos" with line lc 3 , "interact-XIAP-IDE12.pep_FVAL.tsv" using 1:22 >>>> title "Model Neg" with line lc 1 >>>> >>>> ^ >>>> "interact-XIAP-IDE12.pep_FVAL.gp", line 25: warning: Skipping data >>>> file with no valid points >>>> Warning: empty y range [0:0], adjusting to [0:1] >>>> >>>> plot "interact-XIAP-IDE12.pep_PPPROB.tsv" using 2:1 title >>>> "PeptideProphet" with line lt 1 lc 3 , x notitle with line lt 0 lc -1 >>>> >>>> ^ >>>> "interact-XIAP-IDE12.pep_PPPROB.gp", line 17: warning: Skipping data >>>> file with no valid points >>>> >>>> plot "interact-XIAP-IDE12.pep_IPPROB.tsv" using 2:1 title "iProphet" >>>> with line lt 1 lc 3 , "interact-XIAP-IDE12.pep_PPPROB.tsv" using 2:1 title >>>> "PeptideProphet" with line lt 1 lc 1 , x notitle with line lt 0 lc -1 >>>> >>>> ^ >>>> "interact-XIAP-IDE12.pep_IPPROB.gp", line 17: warning: Skipping data >>>> file with no valid points >>>> >>>> plot "interact-XIAP-IDE12.pep_IPPROB.tsv" using 2:1 title "iProphet" >>>> with line lt 1 lc 3 , "interact-XIAP-IDE12.pep_PPPROB.tsv" using 2:1 title >>>> "PeptideProphet" with line lt 1 lc 1 , x notitle with line lt 0 lc -1 >>>> >>>> >>>> ^ >>>> "interact-XIAP-IDE12.pep_IPPROB.gp", line 17: warning: Skipping data >>>> file with no valid points >>>> command completed in 0 sec >>>> >>>> running: "/usr/local/tpp/cgi-bin/PepXMLViewer.cgi -I >>>> /usr/local/tpp/data/projects/lindsey/xiap-ide12/interact-XIAP-IDE12.pep.xml" >>>> Segmentation fault (core dumped) >>>> >>>> command "/usr/local/tpp/cgi-bin/PepXMLViewer.cgi -I >>>> /usr/local/tpp/data/projects/lindsey/xiap-ide12/interact-XIAP-IDE12.pep.xml" >>>> exited with non-zero exit code: 35584 >>>> QUIT - the job is incomplete >>>> >>>> Command FAILED >>>> RETURN CODE:35584 >>>> >>>> -- >>>> You received this message because you are subscribed to the Google >>>> Groups "spctools-discuss" group. >>>> To unsubscribe from this group and stop receiving emails from it, send >>>> an email to [email protected]. >>>> To post to this group, send email to [email protected]. >>>> Visit this group at http://groups.google.com/group/spctools-discuss. >>>> For more options, visit https://groups.google.com/groups/opt_out. >>>> >>> >>> -- >>> You received this message because you are subscribed to the Google >>> Groups "spctools-discuss" group. >>> To unsubscribe from this group and stop receiving emails from it, send >>> an email to [email protected]. >>> To post to this group, send email to [email protected]. >>> Visit this group at http://groups.google.com/group/spctools-discuss. >>> For more options, visit https://groups.google.com/groups/opt_out. >>> >>> -- >>> You received this message because you are subscribed to the Google >>> Groups "spctools-discuss" group. >>> To unsubscribe from this group and stop receiving emails from it, send >>> an email to [email protected]. >>> To post to this group, send email to [email protected]. >>> Visit this group at http://groups.google.com/group/spctools-discuss. >>> For more options, visit https://groups.google.com/groups/opt_out. >>> >> >> -- >> You received this message because you are subscribed to the Google Groups >> "spctools-discuss" group. >> To unsubscribe from this group and stop receiving emails from it, send an >> email to [email protected]. >> To post to this group, send email to [email protected]. >> Visit this group at http://groups.google.com/group/spctools-discuss. >> For more options, visit https://groups.google.com/groups/opt_out. >> > > -- > You received this message because you are subscribed to the Google Groups > "spctools-discuss" group. > To unsubscribe from this group and stop receiving emails from it, send an > email to [email protected]. > To post to this group, send email to [email protected]. > Visit this group at http://groups.google.com/group/spctools-discuss. > For more options, visit https://groups.google.com/groups/opt_out. > -- You received this message because you are subscribed to the Google Groups "spctools-discuss" group. 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