To reconvert or not reconvert?

Spoiler alert: The answer is to reconvert.

In my previous post, I set out a game plan to troubleshoot my pecanpie run. First, Laura and I played around with the .blib file produced as part of my PECAN Run 1 Output.

We followed the instructions in this powerpoint after downloading Skyline daily on the Windows machine. We successfully uploaded the background proteome to Skyline, but when we uploaded the .blib file, we got an error message:

This supports Sean’s finding that nothing was ever generated by PECAN because it couldn’t find what it considered to be valid MS2 data in my mzML files and stopped there. This did not happen to Laura’s files, which is weird because we underwent the same MSConvert process. I will reconvert one raw file to an mzML file and rerun PECAN. Hopefully that will yield a usable .blib file!

The lab notebook I’m working in can be found here. A couple of highlights:

• I had to install ProteoWizard Tools and Git Bash on the Windows computer
• Created a config file for MSConvert
• Used the following code to run MSConvert (which then ensued in the computer crashing the first time, but worked pretty well the second time)

cd C:\Users\srlab\Documents\2017-03-07-MSConvert

"c:\Program Files\ProteoWizard\ProteoWizard 3.0.10577\msconvert.exe" -c C:\Users\srlab\Documents\2017-03-07-MSConvert/msconvert-SIMMS1.config C:\Users\srlab\Documents\2017-03-07-MSConvert/2017_January_23_envtstress_oyster1.raw

• Used the following code for my second PECAN run

pecanpie -o /home/srlab/Documents/DNR_PECAN_Run_2_20170307_Output \
-b /home/srlab/Documents/DNR_PECAN_Run_2_20170307/Combined-gigas-QC.txt \
-n DNR_PECAN_Run_1_20170307_SpLibrary \ 
-s gigas \
--isolationSchemeType BOARDER \
--pecanMemRequest 35 \
/home/srlab/Documents/DNR_PECAN_Run_2_20170307/2017-03-07-mzML-file-path-list.txt \
/home/srlab/Documents/DNR_PECAN_Run_2_20170307/2017-03-07-background-peptides-path-list.txt \
/home/srlab/Documents/DNR_PECAN_Run_2_20170307/2017-03-07-isolation-windows.csv \
--fido \
--jointPercolator

Then I used this code to run the search with pecanpie:

cd [directory specified by the -o argument for pecanpie] \
./run_search.sh

And used qstat -f to check that the job was running.

Tomorrow, I’ll see if the log file has the same error as my first run (and maybe even get a viable .blib file to explore Skyline with while I set up the rest of my analyses……..).