I know someone else got the "Search failed" message before and I had some theories about what might have gone wrong, but I did not hear back so I am not sure if any of those theories were correct:
https://skyline.ms/announcements/home/support/thread.view?rowId=61146

If you would like, you could send us your files and we could see if we can figure out what is going wrong.
You could zip up all your files and upload them here:
https://skyline.ms/files.url

It would also be helpful if you could include some screenshots of the pages of the "Import Peptide Search" wizard so that we can see which settings you used when we try to reproduce your error.

I think in some of these cases the reason that the search failed was that the FASTA file was much too small. I am not sure what the minimum size of the FASTA file should be, but I imagine it should be a few hundred proteins at least.
-- Nick

Hi Nick,

Thanks for responding to my enquiry. I've uploaded the zip file (CalA.d.zip). This should contain the CalA.d (DDA data file), CalA.sky, CalA.fasta, and mzip.pptx (series of screenshots to replicate the error).

PS Please delete the CalA.zip folder as this does not contain the data file.

Thanks in advance!


Kind regards,
Elvis

Hello,
Did you manage to get rid of this error ? Because I'm having th same trouble and indeed my fasta contains only 21 proteins.

Best regards
Victoria

Victoria,

Unfortunately it looks like I never downloaded Elvis's files and investigated this issue. (Sorry about that, Elvis. If you would like, you can upload those files again).

I don't think it makes sense to do a DDA peptide search using a FASTA file with only 21 proteins. Even if you are only interested in a few proteins, things work a lot better if you use a FASTA file for searching that contains many more proteins, perhaps the complete set of proteins you would plausibly expect to find in the sample.

When you are launching a search from the "File > Import > Peptide Search" you are given the opportunity to provide two different FASTA files. The first FASTA file that you are asked to provide is the FASTA file that is used for the search. That should be a sizeable FASTA file. Skyline also allows you to specify a different, often smaller FASTA file which is used to decide which peptides to add to your Skyline document.
-- Nick

Hello Nick,

Thank you for your quick answer!
Actually in my sample I have a mix of 20 peptides so I run a DDA and I wanted to create a library that's why I have my fasta with those 20 peptide. Later I will analyse more complex samples but I'll only focu on the 20 peptides with PRM. So do you know how I can create my library with my DDA run and my fasta ? Thank you for your help.

Best
Victoria

I do not actually know what you are supposed to do if your DDA sample really does only contain 20 peptides and you want to use a peptide search engine to figure out which spectrum belongs to which peptide.

What I imagine that you would do is just run the search engine part of the search and then not run the percolator part.
Then, when you import the peptide search results into Skyline you would specify that you want a 100% FDR, or a 1.0 q-value cutoff or similar.

I am not sure which peptide search engines it is possible to do that with.

Maybe someone else on this support board will be able to give a better answer.
-- Nick