小麦谷蛋白亚基的快速提取分离及SDS-PAGE分析

Rapid separation and SDS-PAGE analysis of wheat glutenin subunits

用70%乙醇和50%异丙醇除去醇溶蛋白,以二硫苏糖醇(DTT)为还原剂,在50%异丙醇,0.08mol/LTris HCl(pH为8.0)缓冲液中提取总麦谷蛋白亚基,加入丙酮至浓度为40%时,高分子量麦谷蛋白亚基(HMW GS)沉淀析出;继续加入丙酮至80%时,析出低分子量麦谷蛋白亚基(LMW GS).在3.3%浓缩胶和10%分离胶的不连续体系中进行SDS PAGE电泳,结果表明,该方法可以有效地除去醇溶蛋白对麦谷蛋白亚基电泳分析的背景干扰,HMW GS和LMW GS的提取、分离一步完成,操作简便,时间短,对大量小麦样品的HMW GS和LMW GS可以快速分离、提纯和鉴定.

After deleting gliadin with 70% ethanol and 50% 2-propanol, the total glutenin subunits are extracted in 50% 2-propanol, 0.08 mol/L Tris-HCl buff, containing 1% DTT. When acetone concentration is added to 40%, high-molecular-weight glutenin subunits (HMW-GS) are precipitated, and acetone concentration is continue added to 80%, purged low-molecular-weight glutenin subunits (LMW-GS) are precipitated. In SDS(-PAGE) electrophoresis (3.3% stacking gel and 10% resolving gel) system, HMW-GS and LMW-GS are gliadin(-free) background. This method can delete gliadin effective and rapid separate HMW-GS and LMW-GS by one step and short time. The method is applicable to wheat protein separation, variety identification and variety modify.