Total protein extraction and 2-D gel electrophoresis methods for Burkholderia species.
作者： Billie Velapatiño , James E. A. Zlosnik , Trevor J. Hird , David P. Speert
DOI: 10.3791/50730
关键词: Two-dimensional gel electrophoresis 、 Lysis 、 Protein purification 、 Polyacrylamide gel electrophoresis 、 Isoelectric point 、 Gel electrophoresis 、 Isoelectric focusing 、 Molecular biology 、 Biology 、 Chromatography 、 Burkholderia
摘要: The investigation of the intracellular protein levels bacterial species is importance to understanding pathogenic mechanisms diseases caused by these organisms. Here we describe a procedure for extraction from Burkholderia based on mechanical lysis using glass beads in presence ethylenediamine tetraacetic acid and phenylmethylsulfonyl fluoride phosphate buffered saline. This method can be used different species, growth conditions, it likely suitable use proteomic studies other bacteria. Following extraction, two-dimensional (2-D) gel electrophoresis technique described study global changes proteomes consists separation proteins according their isoelectric point focusing first dimension, followed basis molecular weight acrylamide second dimension. Visualization separated carried out silver staining.
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