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以Lox缺失体五星1号、3号、4号和非缺失类型大豆中黄13种子为材料,采用非变性聚丙烯酰胺凝胶电泳(Native-PAGE),通过改变分离胶浓度、电泳电压、样品浓度、电泳时间和染色方法等,探索大豆Lox同工酶电泳最佳条件。结果表明:浓缩胶浓度5%,分离胶浓度13%,浓缩胶电泳电压90 V、分离胶电泳电压190 V,样品浓度40%,电泳时间7 h,采用pH8.0、pH6.5两种不同的酶染液结合热考马斯亮蓝染色,4种Lox同工酶(Lox1、Lox2、Lox3a和Lox3b)的分辨清晰度最高。该方法简便安全、经济有效,可用于大豆种质资源Lox缺失体的筛选鉴定。
In this study, native Lox-deficient Wuxing No.1, No.3 and No.4 soybean varieties and non-deletion type soybean Zhonghuang 13 seeds were used as materials. By using native polyacrylamide gel electrophoresis (Native-PAGE) Concentration, electrophoresis time and staining methods to explore the best conditions for soybean Lox isoenzyme electrophoresis. The results showed that the optimal conditions were as follows: concentration of concentrate 5%, concentration of separation gel 13%, gel electrophoresis voltage 90 V, separation gel electrophoresis voltage 190 V, sample concentration 40%, electrophoresis time 7 h, pH8.0, pH6.5 Of the enzyme staining combined with hot Coomassie brilliant blue staining, the highest resolution of the four Lox isoenzymes (Lox1, Lox2, Lox3a and Lox3b). The method is simple, safe and economical, and can be used for screening and identification of Lox deletion bodies in soybean germplasm resources.