家蚕微孢子虫(Nosema bombycis)孢壁蛋白提取方法的优化研究

分别用SDS法、煮沸法、碱溶法和Brosson法,提取家蚕微孢子虫孢壁蛋白,并对孢壁蛋白及处理后的孢子形态进行比较研究。结果表明,不同方法处理后的孢子形态均保持完整,SDS处理后孢子为短杆状且明显变得粗壮,煮沸处理后孢子变小呈梭形,其他方法处理的孢子均有膨胀现象。SDS-PAGE检测显示,SDS法提取的孢壁蛋白出现3条明显的主带,煮沸法提取的孢壁蛋白出现5条明显的主带,0.1 mol/L KOH提取的孢壁蛋白出现2条主带;而Brosson法提取的孢壁蛋白呈现出20~30条带,体现了家蚕微孢子虫孢壁蛋白组成的复杂性,并适合于孢壁蛋白质组学的研究。

Extraction of Spore Wall Proteins of Nosema bombycis With Improved Methods

Comparative study on morphology of Nosema bombycis spores and their wall proteins extracted by different methods were conducted.After treated with different methods,including SDS extraction,boiling extraction,different concentration KOH extraction,K2CO3 extraction and Brosson's extraction,the spores(stained by Giemsa) were in their integrities and changed little under the microscope,but the morphology of spores exhibited some differences.After SDS treatment,the spores became shorter rod in shape but bigger.After boiling,the spores showed smaller and spindle in shape.And treated with the other methods,the spores showed a little expansion.Three main bands of proteins were detected by SDS-PAGE from the spore wall proteins(SWP) extracted by SDS,five main bands from SWP by method of boiling,and two main bands from SWP by 0.1 mol/L KOH extraction.Brosson's extract showed that the spore wall protein bands were around 20～30,suggesting that the composition and function of SWP is very complex and this extract is suitable for proteomics analysis of spore wall proteins.