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An efficient protein preparation for proteomic analysis of developing cotton fibers by 2‐DE
Author(s) -
Yao Yuan,
Yang YiWei,
Liu JinYuan
Publication year - 2006
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.200600111
Subject(s) - phenol extraction , extraction (chemistry) , fiber , chromatography , protein purification , proteome , chemistry , polyphenol , proteomics , tandem mass spectrometry , acetone , biochemistry , mass spectrometry , gene , antioxidant , rna , organic chemistry
Preparation of high‐quality proteins from cotton fiber tissues is difficult due to high endogenous levels of polysaccharides, polyphenols, and other interfering compounds. To establish a routine procedure for the application of proteomic analysis to cotton fiber tissues, a new protocol for protein extraction was developed by optimizing a phenol extraction method combined with methanol/ammonium acetate precipitation. The protein extraction for 2‐DE was remarkably improved by the combination of chemically and physically modified processes including polyvinylpolypyrrolidone (PVPP) addition, acetone cleaning, and SDS replacement. The protocol gave a higher protein yield and vastly greater resolution and spot intensity. The efficiency of this protocol and its feasibility in fiber proteomic study were demonstrated by comparison of the cotton fiber proteomes at two growth stages. Furthermore, ten protein spots changed significantly were identified by MS/tandem MS and their potential relationships to fiber development were discussed. To the best of our knowledge, this is the first time that a protocol for protein extraction from cotton fiber tissues appears to give satisfactory and reproductive 2‐D protein profiles. The protocol is expected to accelerate the process of the proteomic study of cotton fibers and also to be applicable to other recalcitrant plant tissues.