Hello, The first thing I would check is if there are any iTraq signals in the spectra from a few peptide identifications. Click on any "ions" link in the pepXML Viewer, and then click on the "zoom 112-122" checkbox and "GO" buttons on the spectrum viewer that pops up. Can you see any of the reagent peaks? Do this for a few spectra.
The other thing to look for is the correctness of the condition file; how did you generate it? Lastly, were there any errors or warnings in the output of the tools when you ran them? Sometimes it is hard to miss if, say, Libra gave a warning in the midst of all the other output. Let us know what you find, and thanks for trying our tools. --Luis On Fri, Aug 21, 2009 at 10:43 AM, sunsun <[email protected]> wrote: > > This is my first time to use the TPP to process my iTRAQ-labeled > shotgun data, everything went successful until I found the > quantification of the iTRAQ data is [unavailable] in the result sheet > of 'Analyze Peptides', I have no idea why this happened and perhaps > someone here can give me some hint. > Here is the detailed data description and how I load them into the > TPP: > I only used 114 and 117 label from a iTRAQ development kit, just for > method development. Labeled peptides was further separated by SCX > before loaded into RP-HPLC coupled with an Agilent Q-TOF. Spectrum of > each SCX fraction was in .d files and was further converted to mzXML > by Trapper. I also converted .d files to mgf via Agilent's MassHunter > before the search. Database search was completed by Mascot and results > was exported in dat. format, which was further converted to pep.xml in > TPP. I put all pep.xml and corresponding mzXML files in one folder > under the directory of C:\Inetpub\wwwroot\ISB\data. In the TPP, I > selected the Mascot pipeline and loaded all the pep.xml files, > selected Run PeptideProphet and Run PeptideProphet afterwards. Run > libra using the condition file generated according to the isotopic > correction factors come with my kit. Everything else were leave as > default. > The results of running XInteract: both interact.pep.xml and > interact.prot.xml were generated. In the pepXML viewer, column 'pepP > probablity', 'spectrum', 'ionscore', 'identityscore','ions', > 'peptides', 'protein' all look good, but the Libra 114 and 117 columns > both shown [unavailable]. > Any comments, discussions are welcome!! > Thank you guys! > > > > --~--~---------~--~----~------------~-------~--~----~ 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 -~----------~----~----~----~------~----~------~--~---
