汤氏毛藻产杀虫毒素毛藻素A的提取及性质研究

Wei-Zhen Liu, Drion G. Boucias, Clayton W. McCoy
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引用次数: 35

摘要

刘,W.-Z。, Boucias, d.g.和McCoy, c.w. 1995。汤氏毛藻产杀虫毒素毛藻素A的提取及特性研究。真菌学通报,2009,33(4):444 - 444。采用离子交换、凝胶渗透和免疫亲和层析相结合的方法,对由thompsoni Hirsutella var. thompsonii (JAB-04)菌株产生的Hirsutellin A (HtA)进行了提取和纯化。通过氨基酸分析和n端测序证实了纯化的HtA的身份。制备的HtA单克隆抗体可直接夹心ELISA检测25 ~ 50 ng HtA。此外,利用Western blot方法,证明抗体对HtA具有特异性。在深层发酵过程中监测HtA的产生。利用SDS-PAGE凝胶和ELISA对16.3 kda条带进行密度分析,发现细胞外HtA在指数生长后期(39 ~ 45 h)达到峰值(13 ~ 14 μg/ml)。HtA产量与菌丝生长直接相关。21小时培养滤液对大蜡蛾幼虫有剧毒作用。最终浓度为40 pmol的纯HtA对mellonella幼虫具有高度毒性。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Extraction and Characterization of the Insecticidal Toxin Hirsutellin A Produced by Hirsutella thompsonii var. thompsonii

Liu, W.-Z., Boucias, D. G., and McCoy, C. W. 1995. Extraction and characterization of the insecticidal toxin hirsutellin A produced by Hirsutella thompsonii var. thompsonii. Experimental Mycology 19, 254-262. Hirsutellin A (HtA) produced by Hirsutella thompsonii var. thompsonii (strain JAB-04) was extracted and purified using a combination of ion-exchange, gel-permeation, and immunoaffinity chromatography. The identity of the purified HtA was confirmed by amino acid analysis and N-terminal sequencing. Monoclonal antibodies prepared against HtA were capable of detecting 25-50 ng of HtA by direct sandwich ELISA. In addition, utilizing Western blot methods, the antibodies were shown to be specific to HtA. The production of HtA was monitored during submerged fermentation. The peak level of exocellular HtA (13-14 μg/ml) was during the late exponential growth phase (39-45 h), determined by utilizing a combination of densitometric analysis of the 16.3-kDa bands on SDS-PAGE gels and ELISA. HtA production was directly correlated with mycelial growth. Twenty-one-hour culture filtrates were highly toxic to larvae of the greater wax moth. Pure HtA at a final concentration of 40 pmol was highly toxic to Galleria mellonella larvae.

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