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适于SDS-PAGE分析的苹果叶片蛋白质提取方法

DOI: 10.7668/hbnxb.2009.02.016, PP. 75-78

Keywords: 苹果,SDS-PAGE,蛋白质提取

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Abstract:

为探索苹果叶片蛋白质SDS-聚丙烯酰胺凝胶电泳(SDS-PAGE)分析的样品制备方法,比较了三氯乙酸(TCA)/丙酮沉淀法、改良的Tris-HC1法、酚-甲醇/醋酸铵沉淀法、二硫苏糖醇(DTT)/丙酮法及Tris-HC1法等5种蛋白质的提取方法.制备的样品经SDS-PAGE分离采用银染显色.结果表明,改良的Tris-HC1法在加大PVPP用量和蛋白质提取缓冲液的基础上,将丙酮沉淀蛋白质的时间由30min增加到1h,使蛋白质的得率最高为3.23μg/μL,上样量5μL得到的蛋白质条带数量最多,条带最清晰为1条.利用这一改良方法对苹果实生树不同节位蛋白质的动态变化进行了研究,共发现6条蛋白质差异带,分子量分别为71.9,60.5,52.6,1.1,35.3,18.5kDa,条带清楚,背景清晰.因此,改良的Tris-HC1法适用于苹果叶片的SDS-PAGE分析.

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