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spectral library as reference

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spectral library as reference sarah  2013-02-01 06:54
 
Hi Brendan,

I have a quick question concerning use of spectral libraries for results evaluation. If I have several libraries loaded and am looking at an SRM experiment, Skyline will give me a dot product score next to the precursor as well as the order of fragments in both chromatographic peak (from my experiment) and spectrum. Now, when I switch between different libraries (which all contain a spectrum for the peptide of interest), the dotp and order of fragments stays the same although sometimes it's obvious that the spectra are quite different. So I have a feeling that Skyline doesnt update the values here....But then my question would be: Which of the libraries is used when calculating the dotp (and order of fragments)?

Thanks in advance!
Sarah
 
 
Brendan MacLean responded:  2013-02-01 16:05
Hi Sarah,
Skyline is using the spectrum that shows in the MS/MS Spectrum graph by default, and you are right that it does not update these values in the document when you switch which spectrum you are looking at using the dropdown list above the graph.

I believe we still have an open TODO issue to allow the spectrum used to be manually chosen through the toolbar above the graph that appears when there are multiple spectra to choose from. At the moment, however, Skyline always uses the default spectrum, which is chosen from your libraries in the order they appear in the settings. The first library that contains a spectrum in the list is the one that provides the spectrum used for choosing transitions and generating dot-products. You can change the spectrum getting used by changing the order of your libraries, though this may change quite a lot. There is no way yet to change the spectrum manually for just a single peptide.

Hope that helps clarify what is going on. Good luck with your experiments.

--Brendan
 
gabe responded:  2017-09-21 13:16
Hi, I'm wondering if this issue has been addressed? If I have three reference spectral libraries in my Skyline method: A, B, and C - it would be nice to occasionally be able to specify different reference spectra for different peptides. So I could indicate that peptide #1 gets a reference spectrum from library A and peptide #2 gets a reference spectrum from library B, etc.

Can this be done yet?

Thanks

Gabe
 
Brendan MacLean responded:  2017-09-23 16:39
I gave my current answer on this separate new thread:

https://skyline.ms/announcements/home/support/thread.view?rowId=31534