丝状真菌Pythium sp.GY1938菌株蛋白图谱染色方法的比较及优化

Comparison and optimization of staining methods for protein analysis in the Pythium sp.GY1938 strain of filamentous fungus

目的探索制备灭蚊丝状真菌Pythiumsp.GY1938菌株菌丝体可溶性蛋白图谱的最佳染色方案。方法采用不同的破壁方法制备Pythiumsp.GY1938菌丝体可溶性蛋白,并通过SDS-PAGE进行蛋白质电泳分离,进一步采用考马斯亮蓝R-250染色法、Vorum银染法、EMBL银染法、Blum银染法4种染色方案分别进行凝胶染色,对不同染色方案的染色效果进行分析和比较。结果对于Pythiumsp.GY1938菌丝体可溶性蛋白样品中的中、高分子质量蛋白(≥44ku),考马斯亮蓝R-250的染色效果较好,且简单、快捷;对于低分子质量(<44ku)的可溶性蛋白,Vorum银染法染色效果较好;Blum银染法和EMBL银染法的染色效果均不理想。结论考马斯亮蓝R-250染色法和Vorum银染法均可应用于Pythiumsp.GY1938菌丝体可溶性蛋白图谱的制备,前者适合对中、高分子质量蛋白染色,后者适合对低分子质量蛋白染色。

Objective To explore the best methods of staining in order to prepare a map of soluble proteins from the mycelium of the Pythium sp.GY1938 strain of mosquito-killing filamentous fungus.Methods Coomassie brilliant blue staining,Vorum silver staining,EMBL silver staining,and Blum silver staining were used to stain gels and map proteins separately after SDS-PAGE.Results Compared to the three methods of silver staining,Coomassie brilliant blue R-250 staining provided the most effective,simple,and quick staining of soluble proteins with a high molecular weight（HMW） or medium molecular weight（MMW）（≥44 ku）.Vorum silver staining was effective at staining soluble proteins with a low molecular weight（LMW）（44 ku）.Blum and EMBL silver staining were all not effective.Conclusion Both Coomassie brilliant blue R-250 and Vorum silver staining can be used to efficiently map the soluble proteins from the mycelium of the Pythium sp.GY1938 strain.The former is more suitable for mapping medium and high molecular weight proteins while the latter is more suitable for mapping low molecular weight proteins.