Generating mass list

Generating mass list tdpatsch  2022-05-03 06:28

Hi, I am quite new in the field of proteomics and therefore my request might be rather trivial. I have a database containing hypothetical proteins in FASTA format. I would need a program that imports these FASTA sequences, does the tryptic digest in silico and provides me with an MS list that I can use as an inclusion list on a Thermo QExactive Plus to search specifically for these proteins. Somebody told me that skyline can do the job. Would be great if someone could explain how to do it or refer to the proper tutorial.

Nick Shulman responded:  2022-05-03 07:05
Yes, Skyline is a great tool for what you want to do.
You will use the menu item:
File > Import > FASTA
to read in your FASTA file which will add all of the tryptic peptides to your document.

Then, you probably need to go to:
Settings > Transitions Settings > Full Scan
and change MS/MS filtering Acquisition Method to "PRM".
Also, change, the "Retention time filtering" option to "Include all matching scans".
You probably also want to change MS1 filtering isotope peaks included to something other than "None" if you would like Skyline to extract MS1 chromatograms in addition to the PRM chromatograms.

Then, you will export your isolation list using the menu item:
File > Export > Isolation List

After you have collected data on your mass spectrometer you will tell Skyline to extract chromatograms from your raw files using the menu item:
File > Import > Results

There are a lot of tutorials on this page:

Probably, the best tutorial for you to look at is the PRM tutorial:
-- Nick