Frequently Asked Questions
Q: What are the hardware requirements for MSFragger?
A: This depends on the complexity of the search. In general, we recommend at least 8-16 GB, but complex closed searches and open searches will require more. Additional variable modifications, semi-enzymatic, and non-enzymatic/non-specific searches will require additional memory and processing cores to maintain very short search times. (We do not recommend performing an open search with non-specific cleavage rules.) Performance scales well with the number of processors.
Q: Does MSFragger require decoys in the FASTA sequence database?
A: Yes. Philosopher can download and format a database for you (including the addition of reversed/decoy sequences). Custom databases can also be generated and formatted by Philosopher. See this page for more information & usage examples.
Q: How can I convert raw MS/MS files to mzML for MSFragger searches?
A: We recommend using the msconvert tool from ProteoWizard. A tutorial can be found here.
Q: I got a memory error when I tried to run MSFragger. What can I do?
A: First make sure a 64-bit (x64) version of Java/JDK is installed. (The latest version of JDK, which is 64-bit, can be downloaded here.) If you’re using MSFragger in the command line, make sure to specify the appropriate amount of memory given your hardware configuration (e.g.
-Xmx32G to use 32 GB). The database splitting option (available in FragPipe and through the command line) can be used to reduce the size of the in-memory fragment ion index that MSFragger generates. If you still run out of memory, you can reduce the size of the fragment ion index a few ways, 1) use only reviewed sequences in your fasta database, 2) decrease digest_max_length, 3) remove some variable modifications, 4) try a fully-enzymatic search.
Q: Can low resolution (e.g. ion trap) MS/MS data be used in MSFragger?
A: Low resolution MS/MS spectra are suitable for closed searches, but we recommend open searches only be performed on high mass accuracy MS/MS spectra (e.g acquired in an Orbitrap or high-res TOF).