Command line interface for peptide search dsullivan  2019-07-31 11:01
 

Hi Brendan,

Thanks for developing skyline. I am trying to use it to perform some automated analysis for our lab. We have a small set of known peptides and modifications that we want to import and run in skyline.

I have been generating all combinations on my own in python and importing the respective m/z's to skyline as a transition list, however I see that in the recent release there is more support for peptides and the command line (and now included command line documentation). Great!

My questions are:

  1. Is the best way to get my peptides in to import as items in a FASTA? Or should I still use a transition list?
  2. I see I can import a "peptide search results file", but I can't find anywhere that details the format of this file. Can you provide an example?
  3. I see that modifications also go in this search results file, how would that look as well?
  4. There is now an option for charge state, but I don't see an option for specifying my digestion and missed cleavages, does one exist for the command line?

I'm very new to skyline so I apologize if there is an easy resource I have not been able to find.

Regards,
Devin
 
 
Nick Shulman responded:  2019-07-31 13:34
#2: Skyline uses another program called "BiblioSpec" to read peptide search results. This is the list of peptide search result formats that BiblioSpec knows how to read:
https://skyline.ms/wiki/home/software/BiblioSpec/page.view?name=BlibBuild

#3, #4: The commandline allows you to specify a starting Skyline document with the "--in" commandlne parameter. You can use the regular Skyline user interface to create an empty Skyline document with the right digestion and modification settings in it, and save that .sky file.

If you want to learn more about using Skyline with DDA peptide search results, it might be helpful to look at the MS1 tutorial:
https://skyline.ms/wiki/home/software/Skyline/page.view?name=tutorial_ms1_filtering

-- Nick
 
dsullivan responded:  2019-08-01 13:17
Hey Nick,

Thanks for the quick reply.

Hmm, I don't think that BiblioSpec is exactly what I want then. I already know the list of peptides I want to search for and the potential modifications that may be on the peptides. I just want to pass them in and compute proper charge states with/without modifications and find the peaks. I have thus far just been enumerating them in python. My issue is that when I import them I would like skyline to realize they are peptides and add the modifications/charge states etc itself. I think this should yield better results as I see some issues where charge states are now being assigned different elution times because skyline has no association between them. Is there a way to do that?

Yes, I have a dummy input file, but is there no way to reset #'s 3 and 4 via the command line? Ideally I don't want to have to manually create a dummy for each case here since these settings may change quite frequently in our case.


Thanks again for the help!
Regards,
Devin
 
Nick Shulman responded:  2019-08-01 14:21
If you are using the regular Skyline user interface, you can use the menu item:
Edit > Insert > Peptides
to paste in a list of peptide sequences.

Skyline will choose precursors and transitions based on your settings on the Transition Settings Filter tab etc.

I am not sure whether there is any way to get to this peptide inserting functionality via the commandline.

Also, currently there is no way to define modifications from the commandline. We might have some plans to implement this sort of functionality in the future, but I am not sure.
-- Nick