枇杷叶片和果实总蛋白质提取及双向电泳的优化方法

Optimization of protein isolation and method of two-dimensional electrophoresis from leaves and fruit of Loquat(Eriobotrya japonica)

以枇杷叶片和果实为材料,探索适于枇杷总蛋白质分析的双向电泳方法。比较2种不同蛋白质提取方法对双向电泳结果的影响,并对双向电泳蛋白质上样量、等电聚焦条件等进行了优化,探索出一种适于枇杷叶片和果实总蛋白质分析的高分辨率、高重复性双向电泳方法。采用酚抽法提取枇杷叶片和果实蛋白质,以24 cm、pH 4～7的IPG胶条,按150μg蛋白质上样量进行杯上样,在适宜的等电聚焦条件下,可获得蛋白质点数多,背景清晰,重复性好,分辨率高的蛋白质双向电泳图谱。

A better extraction method was established to analyze woody plant protein proteolysis and two-dimensional gel electrophoresis by using Loquat（Eriobotrya japonica） leaves and fruits as experiment materials.Two total protein extraction methods,including trichloroacetic acid/acetone precipitation method and phenol extraction method were compared.We optimized protein sample size and isoelectric focusing section,and designed a good repeatability and high-resolution two dimensional electrophoresis technology to analyze protein in loquat leaves and fruits.We achieved satisfactory 2-DE patterns with well-resolved polypeptide spots throughout the gel by phenol extraction method combination with 150 μg loaded sample amounts.