I will try out with Comet as Damon is doing and see how does it go compared to three searches with XTandem. I also second that, that it would be really useful to have a capable multiplex xpress.
It is also nice to hear from Jimmy that it is doable, and that there's/would be a rewritten xpress, of course with the big caveat to some day integrating it into TPP. If you need some testing with the developing standalone xpress, I will be happy to run some. Best, Alejandro On Friday, May 1, 2015 at 8:44:10 PM UTC+2, Jimmy Eng wrote: > > I can get xpress working with L/M/H SILAC. But I loath the thought of the > work it would take to interface it within the TPP (updating pepXML schema, > xinteract wrapper tool, XPressPeptideRatioParser, XPressProteinRatioParser, > PepXMLViewer, the protein viewer, etc.). > > I will play with my standalone, rewritten xpress (runs much more > efficiently) to get it a bit more functional, including L/M/H SILAC > support, and then maybe we can find another developer to integrate it back > into the TPP. > > > On Thu, Apr 30, 2015 at 10:48 AM, Damon May <[email protected] <javascript:>> > wrote: > >> I have the same issue; I'm running a single search with Comet and then >> PeptideProphet, then copying the output pepXml and running XPress >> separately for H/L and M/L. This gives me answers, but I'd have a lot more >> confidence in an enhanced XPress that was aware of all three labels (in my >> case, light-medium-heavy SILAC) and was choosing scan extents appropriately >> across all the labels. >> >> If anyone's mulling over this kind of enhancement, it would be really >> valuable! >> >> Thanks, >> Damon >> >> >> On Friday, April 24, 2015 at 3:41:02 AM UTC-7, Alejandro wrote: >>> >>> Hi all, >>> >>> Well, so far I've done static searches for each label or doing the >>> variable search as Brian commented, followed by Peptide/ProteinProphet to >>> combine the files, and it somehow works, of course it get's a little bit >>> tricky to handle all of the files. I haven't tried so far used Comet in >>> this setup, I will try and see, looks promising. Regarding the XTandem, I >>> have indeed used it like with several modifications, but I had forgotten >>> that setup, I will try it and see if the ID numbers goes down, compared to >>> doing the three searches. >>> >>> As for the quantification this is where it gets more troublesome. The >>> data that I'm using is Qexactive data, and so far I haven't had so good >>> experience using ASAP ratio, I think we discussed it in a post, was really >>> slow in 4.7, but since TPP 4.8 it is faster, but I see better results using >>> Xpress. So, my strategy so far has been to run it in pairs, do Xpress with >>> Peptide/Protein Prophet to Heavy/Medium and then Heavy/Light and deduce the >>> M/L by dividing the two. The problem so far is in this last step. How to >>> normalize the data (substracting median, etc) when the heavy part is gone. >>> Plus how to handle when one ID is not found in one channel or when the >>> quantification also didn't work in one/two channels. >>> >>> Any ideas? >>> >>> Alejandro >>> >>> >>> >>> On Tuesday, April 21, 2015 at 11:55:58 AM UTC+2, Alejandro wrote: >>>> >>>> Hello all, >>>> >>>> I have some experiments in which the samples were triple dimethyl >>>> labeled. Does any of you have any tips on how to analyze these data with >>>> later version of TPP? >>>> >>>> I looked through the list but only could find a couple of posts. >>>> >>>> So far my ideas would be to do: >>>> >>>> 1. Static searches of Heavy, Medium and Light and then analyze H/M, and >>>> H/L, and deduce the M/L by dividing the ratios of (H/L)/(H/L) ? >>>> >>>> So far I'm doing that, however after Peptide/ProteinProphet the results >>>> of each vary quite abit and might end up with peptides/proteins with a >>>> ratio in just two conditions and not in the three. Usually this also >>>> happens in MaxQuant in triple labeled but only in very few hits. >>>> >>>> 2. Would it be possible to run a variable search for all three >>>> modifications in XTandem in one run, Heavy, Medium, Light and then just >>>> process H/M and H/L with Peptide/ProteinProphet? >>>> >>>> >>>> Maybe some of you had any experience with this, any tips or help would >>>> be appreciated! >>>> >>>> Thanks, >>>> >>>> Alejandro >>>> >>> -- >> 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] <javascript:>. >> To post to this group, send email to [email protected] >> <javascript:>. >> Visit this group at http://groups.google.com/group/spctools-discuss. >> For more options, visit https://groups.google.com/d/optout. >> > > -- 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/d/optout.
