以cotx为分子载体在枯草芽胞杆菌芽胞表面展示绿色-微生物学报.pdfVIP

生 物 工 程 学 报 Chin J Biotech 2010, February 25; 26(2): 264–269 journals.im.ac.cn Chinese Journal of Biotechnology ISSN 1000-3061 cjb@im.ac.cn ©2010 CJB, All rights reserved. 生物技术与方法 以CotX 为分子载体在枯草芽胞杆菌芽胞表面展示绿色 荧光蛋白 1,2 1 1 李倩 ，宁德刚 ，吴春笃 1 江苏大学环境学院，镇江 212013 2 江苏大学生命科学研究院，镇江 212013 摘 要: 芽胞衣壳蛋白CotB 、CotC 、CotG 等可作为芽胞表面展示外源蛋白的分子载体，制备口服重组疫苗或具有催化 活性的重组酶。CotX 为枯草芽胞杆菌Bacillus subtilis 芽胞衣壳中的另一种结构蛋白。为证明CotX 能否作为分子载体 将外源蛋白展示在芽胞表面，本研究将 cotX 基因与绿色荧光蛋白基因 gfp 的编码序列进行基因重组，构建融合表达 CotX-GFP 的整合型重组质粒，将该质粒转化枯草芽胞杆菌，筛选重组菌株并诱导产生芽胞，观察到重组芽胞表面具有 GFP 绿色荧光。结果表明枯草芽胞杆菌的芽胞衣壳蛋白CotX 位于芽胞衣壳外层，可作为芽胞表面展示外源蛋白的载体 分子。 关键词: 枯草芽胞杆菌，芽胞表面展示，CotX ，绿色荧光蛋白 Surface display of GFP using CotX as a molecular vector on Bacillus subtilis spores 1,2 1 1 Qian Li , Degang Ning , and Chundu Wu 1 School of Environment, Jiangsu University, Zhenjiang 212013, China 2 Institute of Life Science, Jiangsu University, Zhenjiang 212013, China Abstract: Spore coat proteins, such as CotB, CotC, CotG et al, are able to efficiently display exogenous protein on spore surface for preparing oral vaccines or enzymes. CotX is another structural protein of spore coats of Bacillus subtilis. To investigate whether CotX could carry target protein onto the spore surface, we constructed a recombinant integrative plasmid, designated as pJS749, which carries a recombinant cotX-gfp gene under the control of the cotX promoter. We transformed pJS749 into Bacillus subtilis 168(trp-), an α-amylase inactivated mutant DRJS749 was selected and confirmed to be a double crossover integrant, where cotX-gfp fragment was integrated into the