Center for Mass Spectrometry and Proteomics | Phone | (612)625-2280 | (612)625-2279
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Proteomics Data Analysis using Galaxy-P
Center for Mass Spectrometry and Proteomics
August 11th 2016
Pratik Jagtap
http://www.cbs.umn.edu/msp
©2016 Regents of the University of Minnesota, All rights reserved.
Center for Mass Spectrometry and Proteomics | Phone | (612)625-2280 | (612)625-2279
© 2015 Regents of the University of Minnesota. All rights reserved.
Documentation: http://z.umn.edu/augworkshopgalaxyp
©2016 Regents of the University of Minnesota, All rights reserved.
Center for Mass Spectrometry and Proteomics | Phone | (612)625-2280 | (612)625-2279
© 2015 Regents of the University of Minnesota. All rights reserved.
GALAXY PLATFORM
Benefits of Galaxy
• A web-based bioinformatics data analysis platform.
• Software accessibility and usability.
• Share-ability of tools, workflows and histories.
• Reproducibility and ability to test and compare results after using multiple
parameters.
• Software tools can be used in a sequential manner to generate analytical workflows
that can be reused, shared and creatively modified for multiple studies.
Goecks J et al Genome Biol. 2010;11(8):R86.
©2016 Regents of the University of Minnesota, All rights reserved.
Center for Mass Spectrometry and Proteomics | Phone | (612)625-2280 | (612)625-2279
© 2015 Regents of the University of Minnesota. All rights reserved.
TOOLS & WORKFLOWS • Software tools can be used in a sequential manner to generate analytical
workflows that can be reused, shared and creatively modified for multiple studies.
For example, Protein Database Downloader
downloads UniProt protein FASTA
databases of various organisms.
©2016 Regents of the University of Minnesota, All rights reserved.
Center for Mass Spectrometry and Proteomics | Phone | (612)625-2280 | (612)625-2279
© 2015 Regents of the University of Minnesota. All rights reserved.
Eng et al 2011 Mol Cell Proteomics. 10(11): R111.009522.
PROTEOMICS WORKFLOW
©2016 Regents of the University of Minnesota, All rights reserved.
Center for Mass Spectrometry and Proteomics | Phone | (612)625-2280 | (612)625-2279
© 2015 Regents of the University of Minnesota. All rights reserved.
WORFLOW 1
INPUTS : PEAKLISTS and SEARCH db
©2016 Regents of the University of Minnesota, All rights reserved.
Center for Mass Spectrometry and Proteomics | Phone | (612)625-2280 | (612)625-2279
© 2015 Regents of the University of Minnesota. All rights reserved.
Tools used in the workflow
©2016 Regents of the University of Minnesota, All rights reserved.
Center for Mass Spectrometry and Proteomics | Phone | (612)625-2280 | (612)625-2279
© 2015 Regents of the University of Minnesota. All rights reserved.
Galaxy-P: https://galaxyp.msi.umn.edu/
©2016 Regents of the University of Minnesota, All rights reserved.
Center for Mass Spectrometry and Proteomics | Phone | (612)625-2280 | (612)625-2279
© 2015 Regents of the University of Minnesota. All rights reserved.
INPUTS : Mass spectral data and search database.
The dataset will be searched against FASTA
database with human proteins, contaminant
proteins, spiked in proteins and a subset of 3-
frame translated cDNA database from
EnSEMBL.
INPUTS: a) MGF formatter MGF files.
(dataset collection)
b) ABRF-Spike4: FASTA sequences of 4
spiked in proteins.
c) FASTA File from EnSEMBL Searches:
Subset of 3-frame translated cDNA database
from EnSEMBL (our template for identifying
novel proteoforms).
d) Human UniProt FASTA file + contaminant
proteins.
HeLa cell lysate
4 proteins spiked in
(10 fmols each)
Digested O/N with trypsin
Liquid chromatography fractionation
(10 fractions)
Thermofinnigan Orbitrap Velos (Orbi MS,
MS/MS HCD)
RAW Files
mzml files
msconvert
MGF files ©2016 Regents of the University of Minnesota, All rights reserved.
Center for Mass Spectrometry and Proteomics | Phone | (612)625-2280 | (612)625-2279
© 2015 Regents of the University of Minnesota. All rights reserved.
RAW DATA CONVERSION TOOL
.RAW
msconvert ProteoWizard
mzML
http://z.umn.edu/msconvert
MGF Formatter
MGF
http://z.umn.edu/mgfformatter
©2016 Regents of the University of Minnesota, All rights reserved.
Center for Mass Spectrometry and Proteomics | Phone | (612)625-2280 | (612)625-2279
© 2015 Regents of the University of Minnesota. All rights reserved.
Log in using your MSI login and password.
Click on http://z.umn.edu/history3
SEARCHING MS/MS SPECTRA A DATABASE
1
2
©2016 Regents of the University of Minnesota, All rights reserved.
Center for Mass Spectrometry and Proteomics | Phone | (612)625-2280 | (612)625-2279
© 2015 Regents of the University of Minnesota. All rights reserved.
Eng et al 2011 Mol Cell Proteomics. 10(11): R111.009522.
PROTEOMICS WORKFLOW
©2016 Regents of the University of Minnesota, All rights reserved.
Center for Mass Spectrometry and Proteomics | Phone | (612)625-2280 | (612)625-2279
© 2015 Regents of the University of Minnesota. All rights reserved.
INPUTS : Mass spectral data and search database.
The dataset will be searched against FASTA
database with human proteins, contaminant
proteins, spiked in proteins and a subset of 3-
frame translated cDNA database from
EnSEMBL.
INPUTS: a) MGF formatter MGF files.
(dataset collection)
b) ABRF-Spike4: FASTA sequences of 4
spiked in proteins.
c) FASTA File from EnSEMBL Searches:
Subset of 3-frame translated cDNA database
from EnSEMBL (our template for identifying
novel proteoforms).
d) Human UniProt FASTA file + contaminant
proteins.
HeLa cell lysate
4 proteins spiked in
(10 fmols each)
Digested O/N with trypsin
Liquid chromatography fractionation
(10 fractions)
Thermofinnigan Orbitrap Velos (Orbi MS,
MS/MS HCD)
RAW Files
mzml files
msconvert
©2016 Regents of the University of Minnesota, All rights reserved.
Center for Mass Spectrometry and Proteomics | Phone | (612)625-2280 | (612)625-2279
© 2015 Regents of the University of Minnesota. All rights reserved.
Select History 1
Import history
Start using this history
Select Workflow 1
Import workflow
Start using this workflow
Run Workflow 1
INPUT
WORKFLOW
©2016 Regents of the University of Minnesota, All rights reserved.
Center for Mass Spectrometry and Proteomics | Phone | (612)625-2280 | (612)625-2279
© 2015 Regents of the University of Minnesota. All rights reserved.
A face in the crowd: recognizing
peptides through
database search.
Eng et al 2011 Mol Cell
Proteomics. 10(11)
PROTEOMICS WORKFLOW
©2016 Regents of the University of Minnesota, All rights reserved.
Center for Mass Spectrometry and Proteomics | Phone | (612)625-2280 | (612)625-2279
© 2015 Regents of the University of Minnesota. All rights reserved.
Visualizing parameters for SearchGUI analysis
©2016 Regents of the University of Minnesota, All rights reserved.
Center for Mass Spectrometry and Proteomics | Phone | (612)625-2280 | (612)625-2279
© 2015 Regents of the University of Minnesota. All rights reserved.
SEARCHGUI
Vaudel M. et al Proteomics (2011) 11(5)
https://code.google.com/p/searchgui/
©2016 Regents of the University of Minnesota, All rights reserved.
Center for Mass Spectrometry and Proteomics | Phone | (612)625-2280 | (612)625-2279
© 2015 Regents of the University of Minnesota. All rights reserved.
PEPTIDESHAKER
Vaudel et al Nature Biotechnology, 33, (2015)
http://galaxyproteomics.github.io/peptideshaker/
©2016 Regents of the University of Minnesota, All rights reserved.
Center for Mass Spectrometry and Proteomics | Phone | (612)625-2280 | (612)625-2279
© 2015 Regents of the University of Minnesota. All rights reserved.
4.3 Peptide Shaker in GalaxyP
©2016 Regents of the University of Minnesota, All rights reserved.
Center for Mass Spectrometry and Proteomics | Phone | (612)625-2280 | (612)625-2279
© 2015 Regents of the University of Minnesota. All rights reserved.
PEPTIDESHAKER: OUTPUTS
©2016 Regents of the University of Minnesota, All rights reserved.
Center for Mass Spectrometry and Proteomics | Phone | (612)625-2280 | (612)625-2279
© 2015 Regents of the University of Minnesota. All rights reserved.
PEPTIDESHAKER: OUTPUTS
©2016 Regents of the University of Minnesota, All rights reserved.
Center for Mass Spectrometry and Proteomics | Phone | (612)625-2280 | (612)625-2279
© 2015 Regents of the University of Minnesota. All rights reserved.
http://z.umn.edu/augworkshopgalaxyp
©2016 Regents of the University of Minnesota, All rights reserved.
Center for Mass Spectrometry and Proteomics | Phone | (612)625-2280 | (612)625-2279
© 2015 Regents of the University of Minnesota. All rights reserved.
QUESTIONS?
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Visit
http://galaxyp.msi.umn.edu
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