Top down / bottom up, MS/MS analysis tool for DDA and DIA mass spectrometry data
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README.md

Informed Proteomics

The Informed Proteomics project includes algorithms for proteomic mass spectrometry data analysis. Although the back-end data access and some of the scoring routines are general purpose, this repository is currently maintained for top down MS/MS datasets.

Install/Tutorials

See the Informed Proteomics GitHub wiki for usage, tutorials, and more!

Downloads

https://github.com/PNNL-Comp-Mass-Spec/Informed-Proteomics/releases

Continuous Integration

The latest versions of the Informed Proteomics tools are available on the AppVeyor CI server

Build status

MSPathFinderT

MSPathFinder finds peptides in top-down LC-MS/MS datasets. Similar to database search engines for bottom-up, it takes a fasta file, a spectrum file, and a list of modifications as an input and reports proteoform spectrum matches (PsSMs) and their scores. These results are output in a tab-separated format and in a MzIdentML file.

Processing steps:

  1. Run PbfGen.exe to convert the instrument file or .mzML file to an optimized binary file
  • Creates a .pbf file This file contains spectra information and full chromatograms for MS1 and MSn data, allowing fast access to extracted ion chromatograms during the search.
  • This step usually only needs to be performed once for a dataset, and by default will do nothing if it has previously been run on the dataset and can find the file.
  • ProMex and MSPathFinderT will perform this step automatically if they are given a spectrum file that is not a .pbf file.
  1. Run ProMex on the .Pbf file to deisotope the data, including determine charge states
  • Creates a .ms1ft file
  • This file can be reused for multiple searches, as long as none of its parameters change
  • MSPathFinderT will perform this step automatically if not provided with a path to a feature file, using the respective parameters from the MSPathFinderT parameters.
  1. Run MSPathfinderT with the .Pbf (or .raw) file and a fasta file to search for proteins
  • Creates _IcTda.tsv files and .mzid
  • Can also be run with the spectrum source file directly, with PbfGen and ProMex run as part of the process.

Example command lines:

PbfGen.exe -s MyDataset.raw

ProMex.exe -i MyDataset.pbf -minCharge 2 -maxCharge 60 -minMass 3000 -maxMass 50000 -score n -csv n -maxThreads 0

MSPathFinderT.exe -s MyDataset.pbf -feature MyDataset.ms1ft -d C:\FASTA\ProteinList.fasta -o C:\WorkFolder -t 10 -f 10 -m 1 -tda 1 -minLength 21 -maxLength 300 -minCharge 2 -maxCharge 30 -minFragCharge 1 -maxFragCharge 15 -minMass 3000 -maxMass 50000 -mod MSPathFinder_Mods.txt

Results viewer / GUI

For viewing search results, you might want to consider LCMS-Spectator. It can also function as a GUI for running ProMex and MSPathFinder.

ProMex Syntax

ProMex version 1.0.6232 (January 23, 2017)
Usage: ProMex.exe
        [-i InputFolder or InputFile]
        [-o OutFolder (default : InputFolder)]
        [-minCharge MinCharge] (minimum charge state, default: 1)
        [-maxCharge MaxCharge] (maximum charge state, default: 60)
        [-minMass MinMassInDa] (minimum mass in Da, default: 2000.0)
        [-maxMass MaxMassInDa] (maximum mass in Da, default: 50000.0)
        [-featureMap y or n (default: y)]
        [-score y or n (default: n)]
        [-maxThreads 0 (default: 0 (automatic set))]

Syntax to create a PNG of the features in an existing ms1ft file
(requires both a .pbf file and a .ms1ft file)
ProMex.exe
        [-i PbfFile]
        [-o OutFolder (default : InputFolder)]
        [-minMass MinMassInDa] (minimum mass in Da, default: 2000.0)
        [-maxMass MaxMassInDa] (maximum mass in Da, default: 50000.0)
        [-ms1ft FeaturesFilePath (use a period to infer the name from the pbf file)]

PbfGen Syntax

PbfGen version 1.0.6232 (January 23, 2017)
Usage: PbfGen.exe
        -s RawFilePath (*.raw or directory)
        [-o OutputDir]

MSPathFinder Syntax

MSPathFinderT version 1.0.6232 (January 23, 2017)
Usage: MSPathFinderT.exe
        -s SpectrumFile (*.raw)
        -d DatabaseFile (*.fasta or *.fa)
        [-o OutputFolder]
        [-m SearchMode] (0: multiple internal cleavages, 1: single internal cleavage (default), 2: no internal cleavage)
        [-tagSearch 0/1] (1: include tag-based search (default), 0: skip tag-based search)
        [-mod ModificationFileName] (modification file, default: no modification)
        [-t PrecursorToleranceInPpm] (e.g. 10, Default: 10)
        [-f FragmentIonToleranceInPpm] (e.g. 10, Default: 10)
        [-tda 0/1] (0: don't search decoy database (default), 1: search shuffled decoy database)
        [-minLength MinSequenceLength] (minimum sequence length, default: 21)
        [-maxLength MaxSequenceLength] (maximum sequence length, default: 500)
        [-minCharge MinPrecursorCharge] (minimum precursor ion charge, default: 2)
        [-maxCharge MaxPrecursorCharge] (maximum precursor ion charge, default: 50)
        [-minFragCharge MinPrecursorCharge] (minimum fragment ion charge, default: 1)
        [-maxFragCharge MaxPrecursorCharge] (maximum fragment ion charge, default: 20)
        [-minMass MinSequenceMassInDa] (minimum sequence mass in Da, default: 3000.0)
        [-maxMass MaxSequenceMassInDa] (maximum sequence mass in Da, default: 50000.0)
        [-feature FeatureFile] (*.ms1ft, *_isos.csv, or *.msalign, default: Run ProMex)
        [-threads MaxNumThreads] (maximum number of threads, default: 0 automatic setting)
        [-scansFile FilePath] (text file with MS2 scans to process)
        [-act ActivationMethod] (0: CID, 1: ETD, 2: HCD, 3: ECD, 4: PQD, 5: UVPD, 6: Unknown/as written

Enabling tag-based searching with -tagSearch 1 can give 5% to 10% more matches, but can increase the runtime by 30% to 50%.

Supported file formats

These programs, when used with no other software installed, only support the use of centroid mzML files as spectrum input. If Thermo Finnigan MSFileReader is installed, it also supports reading from Thermo Finnigan .raw files (Download here, requires registration to download).

Several other formats are supported if an appropriate version of ProteoWizard is installed (Download here, make sure the version downloaded matches system architecture)

MSPathFinder Mods File

See the Example_Files folder for sample modification definition files.

System Requirements

Minimum required:

  • .NET 4.5.1 installed
  • Will probably try to run on anything where you can install .NET 4.5.1.

Minimum recommended:

  • 2.4 GHz, quad-core CPU
  • 16 GB RAM
  • Windows 7 or newer
  • 250 GB hard drive

Written by Sangtae Kim, Junkap Park, and Chris Wilkins for the Department of Energy (PNNL, Richland, WA) Copyright 2015, Battelle Memorial Institute. All Rights Reserved.

E-mail: matthew.monroe@pnnl.gov or proteomics@pnnl.gov Website: http://panomics.pnnl.gov/ or http://omics.pnl.gov


Licensed under the Apache License, Version 2.0; you may not use this file except in compliance with the License. You may obtain a copy of the License at http://www.apache.org/licenses/LICENSE-2.0


Notice: This computer software was prepared by Battelle Memorial Institute, hereinafter the Contractor, under Contract No. DE-AC05-76RL0 1830 with the Department of Energy (DOE). All rights in the computer software are reserved by DOE on behalf of the United States Government and the Contractor as provided in the Contract. NEITHER THE GOVERNMENT NOR THE CONTRACTOR MAKES ANY WARRANTY, EXPRESS OR IMPLIED, OR ASSUMES ANY LIABILITY FOR THE USE OF THIS SOFTWARE. This notice including this sentence must appear on any copies of this computer software.