Protein identification and quantification from riverbank grape, Vitis riparia: Comparing SDS-PAGE and FASP-GPF techniques for shotgun proteomic analysis.
Protein sample preparation optimisation is critical for establishing reproducible high throughput proteomic analysis. In this study, two different fractionation sample preparation techniques (in-gel digestion and in-solution digestion) for shotgun proteomics were used to quantitatively compare proteins identified in Vitis riparia leaf samples. The total number of proteins and peptides identified were compared between filter aided sample preparation (FASP) coupled with gas phase fractionation (GPF) and SDS-PAGE methods. There was a 24% increase in the total number of reproducibly identified proteins when FASP-GPF was used. FASP-GPF is more reproducible, less expensive and a better method than SDS-PAGE for shotgun proteomics of grapevine samples as it significantly increases protein identification across biological replicates. Total peptide and protein information from the two fractionation techniques is available in PRIDE with the identifier PXD001399 (http://proteomecentral.proteomexchange.org/dataset/PXD001399).
Department of Chemistry and Biomolecular Sciences, Macquarie University, North Ryde, NSW, Australia.
Iniga S
IS
George
Plant Science Department, South Dakota State University, Brookings, SD, USA.
Anne Y
AY
Fennell
Department of Chemistry and Biomolecular Sciences, Macquarie University, North Ryde, NSW, Australia.
Paul A
PA
Haynes
D005591:Chemical Fractionation / Q000379:methods*; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D010455:Peptides / Q000032:analysis / Q000737:chemistry; D010940:Plant Proteins / Q000032:analysis* / Q000378:metabolism; D040901:Proteomics / Q000379:methods*; D015203:Reproducibility of Results; D027843:Vitis / Q000737:chemistry* / Q000378:metabolism
D010455:Peptides; D010940:Plant Proteins
Filter Aided Sample Preparation (FASP); Gas Phase Fractionation (GPF); Grapevine; Label-free quantitative shotgun proteomics; Plant proteomics; Vitis riparia