Vagisha Sharma responded: |
2016-09-30 09:00:18 |
Hi Allison,
Do you have a spectrum library associated with your Skyline document? You can check under Settings > Peptide Settings > Library. When I open the attached Weekly_yeast_QEx2.sky I don't see any spectrum libraries selected in settings. But two of the peptides (IATYQER, TASGNIIPSSTGAAK) should have associated spectrum library information, as per the .sky file. I wonder how that happened. Would you please send me a shared zip file from Skyline - from the "File" menu select "Share..". If you are asked to choose between "complete" and "minimal" please choose "complete".
Thanks,
Vagisha |
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allikrest responded: |
2016-10-03 06:44:25 |
Hi Vagisha,
I ended up deleting the document and making a new one, and that seemed to solve the problem! However, now I am having a different problem! Three of the peptides in a different skyline document aren't showing up (see screen shot below and attachment). There is no error message this time. Is this due to the same type problem? I have since created three other AutoQC subfolders with their own skyline files for our different instruments and have not run into this problem, so I am not sure what is going on with this particular file.
Thanks,
Allison |
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Vagisha Sharma responded: |
2016-10-03 12:44:15 |
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allikrest responded: |
2016-10-03 13:15:21 |
Hi Vagisha,
Here is the screen shot again, sorry I must have not attached it previously. All 11 precursors show up in the skyline file but 3 of them don't show up in Panorama for most of the runs (for some reason there are three files/runs that do show them on Panorama).
Allison |
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Vagisha Sharma responded: |
2016-10-03 15:21:35 |
Hi Allison,
Thanks for the screenshot. I had a look at the Skyline documents uploaded to this folder, and only the last three or so have chromatograms for the three peptides (YICDNQDTISSK, LVTDLTK, LVNELTEFAK) in all the replicates. The older Skyline doucments don't have any chromatograms for these peptides.
For example, Weekly_BSA_QEx_2016-09-29_14-47-51, uploaded on 9/29 did not have any data for these peptides from the raw file BSA_40fmol_02_09-12-16_QEx. But there is data for same raw file in the document uploaded on 9/30.
Do you know how that happened? Did you add these peptides to your Skyline documents after you had done the initial raw data import? And then did you re-import the raw files sometime on 9/30 after the document had already been uploaded to Panorama?
Panorama imports data from a replicate (raw file) only once. So, for example, data from BSA_40fmol_02_09-12-16_QEx, with no chromatograms for the three peptides, was imported on 9/29. Data from this replicate was ignored from all Skyline documents uploaded after that.
You could, at this point, delete all the older uploads (see under the "Runs" tab in your folder) and re-upload the latest version of your Skyline document to Panorama. This will import data from all the replicates that are getting ignored right now.
I hope this helps.
-Vagisha |
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allikrest responded: |
2016-10-03 15:45:36 |
Vagisha,
Thank you so much. Your guess at the scenario that caused this is exactly correct! I did alter the peptides in my skyline document after the initial import. I was seeing a message in the AutoQC Loader log about certain relocates being ignored and couldn't figure out why but this all makes perfect sense now. When you refer to re-uploading my latest skyline document, do you mean that I should do it manually? Should I also manually the delete the old document or will it be ignored if a newer version is uploaded? Once again, thank you so much for your help!
Allison |
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Vagisha Sharma responded: |
2016-10-03 16:26:56 |
Hi Allison,
You don't need to manually upload your Skyline document if you are using AutoQC Loader. It will get uploaded next time a new raw file is added. You will, however, have to manually delete the old documents (from the "Runs" tab in your folder in PanoramaWeb"). This is required so that all the data associated with replicates in those documents is removed from the Panorama database, and when AutoQC uploads the document again data for all the replicates that were just deleted will get re-imported and this time it will include your three peptides. Sorry for the inconvenience.
Let me know if you run into any issues.
Thanks!
-Vagisha |
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