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Quantitative proteomics using uniform 15 N‐labeling, MASCOT, and the trans‐proteomic pipeline
Author(s) -
Palmblad Magnus,
Bindschedler Laurence V.,
Cramer Rainer
Publication year - 2007
Publication title -
proteomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.26
H-Index - 167
eISSN - 1615-9861
pISSN - 1615-9853
DOI - 10.1002/pmic.200700180
Subject(s) - mascot , pipeline (software) , proteomics , quantitative proteomics , mass spectrometry , computational biology , computer science , workflow , software , label free quantification , chemistry , chromatography , biology , biochemistry , database , gene , political science , law , programming language
Stable isotope labeling combined with MS is a powerful method for measuring relative protein abundances, for instance, by differential metabolic labeling of some or all amino acids with 14 N and 15 N in cell culture or hydroponic media. These and most other types of quantitative proteomics experiments using high‐throughput technologies, such as LC‐MS/MS, generate large amounts of raw MS data. This data needs to be processed efficiently and automatically, from the mass spectrometer to statistically evaluated protein identifications and abundance ratios. This paper describes in detail an approach to the automated analysis of uniformly 14 N/ 15 N‐labeled proteins using MASCOT peptide identification in conjunction with the trans‐proteomic pipeline (TPP) and a few scripts to integrate the analysis workflow. Two large proteomic datasets from uniformly labeled Arabidopsis thaliana were used to illustrate the analysis pipeline. The pipeline can be fully automated and uses only common or freely available software.