Biology Reference
In-Depth Information
Fig.
2
Pipeline based on freely available tools for spectral counting proteomics. In the present figure we suggest
the GeLC approach as a prefractionation strategy. The bioinformatics steps described herein are bold high-
lighted and span from SEQUEST result file conversion to cluster analysis
2. Install Ruby latest version and be sure to check the options
“Add Ruby executables to your PATH” and “Associate .rb
and .rbw file with this Ruby installation”.
3. Install mspire-sequest by starting command prompt and run-
ning “gem install mspire-sequest”.
4. For conversions, run the executable “srf_to_sqt.rb”. The con-
verter will work on any number of SRF files as input. A simple way
to convert a batch of files is to copy-paste all SRF files in a direc-
tory and run the executable “srf_to_sqt.rb directoryname\*.srf”.
Search Processor Engine (SEPro) is a tool developed to filter PSMs
proposed by the SEQUEST search engine and improve the num-
ber of peptide and protein identifications based on a sophisticated
algorithm. For details on the calculation of the Bayesian score and
how SEPro uses a three-tier approach to optimize the number of
identifications, please refer to Carvalho and coworkers [
28
].
3.2 Peptide
Spectrum Match
Filtering
1. Group all SQTs from each biological replicate into a single
directory. Spectrum processing is carried out per replicate
(i.e., per biological sample).
