两株苏云金芽胞杆菌晶体与芽胞形成过程的细胞学对比分析  

Comparative Cytological Analysis of the Parasporal Crystal and Spore Formation Process in two Bacillus thuringiensis Strains

作　　者：全梅芳[1,2] 彭金莉 何昊城 夏立秋 丁学知[1,2] QUAN Meifang;PENG Jinli;HE Haocheng;XIA Liqiu;DING Xuezhi(School of Medicine,Hunan Normal University,Changsha 410081,China;Hunan Provincial Key Laboratory of Microbial Molecular Biology,State Key Laboratory of Developmental Biology of Freshwater Fish,College of Life Science,Hunan Normal University,Changsha 410081,China)

机构地区：[1]湖南师范大学医学院,长沙410013 [2]微生物分子生物学湖南省重点实验室,淡水鱼类发育生物学国家重点实验室,湖南师范大学生命科学学院,长沙410081

出　　处：《激光生物学报》2020年第2期135-141,共7页Acta Laser Biology Sinica

基　　金：国家自然科学基金面上项目(31370116,31770106);长沙市科技计划项目(kq1907130);湖南省重点科技计划项目(2019NK2192)。

摘　　要：苏云金芽胞杆菌(Bt)中绝大多数杀虫晶体蛋白(ICPs)的表达依赖于芽胞形成,为了从细胞水平研究晶体与芽胞形成之间的关系,本文选用Bt 4.0718与工程菌BtΔleuB为研究对象,利用FM4-64对不同生长阶段的菌体细胞染色,并用激光共聚焦扫描显微镜进行了对比观察和分析。结果显示,Bt 4.0718芽胞的发育依次经历了不对称隔膜和内吞形态学阶段后,能顺利进入下一个发育阶段,直至完成芽胞发育过程后母细胞凋亡裂解;而BtΔleuB细胞进入不对称隔膜期的时间明显延迟,且芽胞发育被阻滞于内吞阶段,伴胞晶体形成最早于不对称隔膜期可见,并且晶体体积继续增大直至细胞死亡。qRT-PCR结果显示,σE、spoIIR和spoIIGA的高水平转录是维持BtΔleuB细胞中ICPs正常表达的关键因素。本研究结果对进一步揭示晶体与芽胞形成之间的关系及构建性状优良Bt工程菌具有一定的参考价值。The expression of most insecticidal crystal proteins(ICPs)in Bacillus thuringiensis(Bt)is dependent on spore formation.In order to study the relationship between ICPs expression and spore formation at the cellular level,a pair of strains Bt 4.0718 and BtΔleuB were selected and subjected to FM4-64 staining at different growth stages for a laser confocal microscope observation.Results showed that the Bt 4.0718 strain underwent the asymmetric septum and forespore engulfment stages successively,and could enter into the next stage and successfully complete the entire process of spore development before the mother cell apoptosis.The asymmetric septum stage of BtΔleuB was significantly delayed,and the spore development was arrested in forespore engulfment.The crystals were visible since the asymmetric cell division stage,and the volume continued to increase until the mother cell died.qRT-PCR result revealed that the high transcriptional level ofσE,spoIIR and spoIIGA is a key factor to maintain the normal expression of ICPs in BtΔleuB cells.These results possessed a certain reference value for further revealing the relationship between sporulation and ICPs expression,and construction of engineering strains with better behavior.

关 键 词：苏云金芽胞杆菌 杀虫晶体蛋白 芽胞 σE因子 

分 类 号：Q939.96[生物学—微生物学]