一种快速的水稻不同组织总蛋白提取方法

A Simple and Rapid Applicable Protocol for Extracting High-quality Total Proteins from Rice

介绍一种简单快捷的水稻不同组织总蛋白提取方法。该方法采用预冷甲醇和丙酮依次洗涤去除杂质和次生代谢干扰物,同时在甲醇洗涤步骤中加入蛋白酶抑制剂混合液,能够最大限度地包含所有蛋白质并降低蛋白酶的降解作用,提高蛋白质的质量和产量。采用该方法从水稻的根、茎、叶及发育中的种子中提取粗蛋白,产量为20～99mg·g-1FW,提取蛋白整个过程耗时1.6h。以SDS-PAGE胶及Western Blot检测所提总蛋白的质量,分别用核蛋白、胞浆蛋白、膜蛋白以及淀粉结合型蛋白等不同类型蛋白抗体进行Western Blot检验。结果显示采用该方法所分离的总蛋白质量可以满足水稻不同类型蛋白对Western Blot的质量要求。

A simple and rapid applicable protocol for extracting high-quality proteins from rice tissues is described . In the protocol , precooling methanol and acetone is used to precipitate proteins and remove contaminants and secondary metabolites . Adding protease inhibitors in methanol for washing is include all proteins to maximize and reduce degradation of proteases .By using this protocol the total protein yield of different tissues were range from 20 to 99 mg ？ g - 1 FW and the entire process taken 1.6 h . To evaluate the quality of the extracted protein ,SDS/PAGE gel and Western Blot were employed . 4 antibodys represented different protein type include the nucleoprotein antibody ,plasmosin antibody , membrane protein antibody and granule-bound starch protein antibody as first antibody for Western Blot . The results show the extraction of high-quality protein samples suitable for Western analysis ,and the overall quality of the protein profiles was good . The simplified protocol described here presents considerable savings of time and reagents .