Hello Shagun, Thank you! I was able to download your data this time and convert it to mzML files. Internally, we use tdf2mzml.py to convert the .d directories into mzML files for the TPP for DIA data, (diapysef for DDA data.) Here is an example of the command we use on our linux computer:
tdf2mzml.py --ms1_type centroid --compression zlib -i A_500ng_DDA_HM10_100_1p9_25minGT_Slot2-1_1_4647.d -o A_500ng_DDA_HM10_100_1p9_25minGT_Slot2-1_1_4647.mzML Would you be willing to try using that converter to generate the mzML files before you search with Comet+TPP? Thanks! -David On Tue, Aug 13, 2024 at 2:16 PM Shagun Gupta <[email protected]> wrote: > Hi David > > Apologies for the hassle! I have uploaded the .d files in an unzipped > format under TPP_diagnosis/TPP_diagnosis. Let me know if this works instead? > > Best > Shagun > > On Tuesday, August 13, 2024 at 3:09:37 PM UTC-4 David Shteynberg wrote: > >> Hello Shagun, >> >> I downloaded the larger zip file twice and tried to decompress, but each >> time it told me there was a corruption in the zip file. The smaller files >> downloaded fine. Can you check your file and upload again? >> >> Thanks! >> -David >> >> On Tue, Aug 13, 2024 at 9:47 AM Shagun Gupta <[email protected]> wrote: >> >>> Hi David >>> >>> I have attached a link to the following files >>> - one replicate per condition .d file (file starting with "C_" had IDs >>> but no values that could be extracted for quantification) >>> - comet parameter file I used >>> - fasta file I ran the search with. >>> >>> Let me know if I can share anything else! >>> >>> Box_link_diagnosis >>> <https://cornell.box.com/s/5y2uak2twszajaniaqpnevxn4zdneu54> >>> Best >>> Shagun >>> >>> On Tuesday, August 13, 2024 at 11:05:42 AM UTC-4 David Shteynberg wrote: >>> >>>> Hello Shagun, >>>> >>>> Thank you for the detailed report. If you are able, please first >>>> compress (into a zip or similar) and then share some of the problem .d >>>> files so I can try to replicate this issue on my computer before I offer >>>> any suggestions. >>>> >>>> Cheers! >>>> -David >>>> >>>> On Tue, Aug 13, 2024 at 7:50 AM Shagun Gupta <[email protected]> >>>> wrote: >>>> >>>>> Hi all >>>>> >>>>> I have been experiencing issues processing .d files obtained from a >>>>> Bruker timsTOF HT in DDA-LFQ mode, more specifically getting >>>>> quantification >>>>> - precursor intensity - per spectra. I am using TPP V6.3.3 Arcus on a >>>>> windows computer. >>>>> >>>>> Details >>>>> - There are 12 .d files (4 repeats of 3 conditions) composed of a >>>>> human with yeast proteome spike in at different ratios. >>>>> - Converted to .mzXML using msconvert >>>>> - Searched with COMET and a search database taken from UniProt for >>>>> Homo sapiens+Yeast >>>>> - Processed with PeptideProphet (filtered at probability associated >>>>> with 1% FDR), XPRESS, ProteinProphet. Ran with -PREC flag >>>>> (PeptideProphet), >>>>> -i flag (XPRESS) >>>>> - Want to do hypothesis testing (comparing the three conditions >>>>> pairwise) using MSstats. So require raw precursor intensity values to make >>>>> a file that can be used as input to MSstats. >>>>> >>>>> Unfortunately after trying the above, and a few more things, while I >>>>> get a large number of PSMs passing FDR (~30k), a large proportion of them >>>>> do not have any precursor intensity value (<1k have some "light area" >>>>> values). Using the "light area" values also does not give expected results >>>>> (its a benchmarking dataset and processing with MSFragger gave excellent >>>>> results that align with expected ratios etc.). Could you suggest things I >>>>> could be doing differently to get the right results? (I imagine it might >>>>> have something to do with the initial conversion to mzXML itself?) >>>>> >>>>> Happy to share any other details needed! >>>>> >>>>> Best >>>>> Shagun >>>>> >>>>> -- >>>>> 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 view this discussion on the web visit >>>>> https://groups.google.com/d/msgid/spctools-discuss/cf666974-5c1f-4de0-80ad-d9b1df2173dan%40googlegroups.com >>>>> <https://groups.google.com/d/msgid/spctools-discuss/cf666974-5c1f-4de0-80ad-d9b1df2173dan%40googlegroups.com?utm_medium=email&utm_source=footer> >>>>> . >>>>> >>>> -- >>> 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 view this discussion on the web visit >>> https://groups.google.com/d/msgid/spctools-discuss/21faed8c-1c4d-45b8-a281-f3f572864ed1n%40googlegroups.com >>> <https://groups.google.com/d/msgid/spctools-discuss/21faed8c-1c4d-45b8-a281-f3f572864ed1n%40googlegroups.com?utm_medium=email&utm_source=footer> >>> . >>> >> -- > 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 view this discussion on the web visit > https://groups.google.com/d/msgid/spctools-discuss/1e530c4b-d879-4117-9708-ae204d2198e5n%40googlegroups.com > <https://groups.google.com/d/msgid/spctools-discuss/1e530c4b-d879-4117-9708-ae204d2198e5n%40googlegroups.com?utm_medium=email&utm_source=footer> > . > -- You received this message because you are subscribed to the Google Groups "spctools-discuss" group. 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