摘要
为了证明苏云金芽胞杆菌以色列亚种２０ｋＤｅ蛋白质对ＣｙｔＡ蛋白溶细胞作用的影响， 根据２０ｋＤｅ蛋白质和ｃｙｔＡ蛋白基因的核苷酸序列，用ＡＭＰＬＩＦＹ程序设计了一套带有酶切位 点的引物，经ＰＣＲ扩增分别获得了２０ｋＤｅ蛋白质和ｃｙｔＡ蛋白基因。将其基因与表达载体 ｐＵＨＥ２４连接并转化到大肠杆菌ＸＬＩ和ＤＨＳ 分别获得含２０ｋＤａ蛋白质基因的克隆子 ＬＺ２９；含ｃｙｔＡ基因的克隆子ＬＺｃｙｔＡ和含有二者基因的重组子ＬＺ２０Ａ．在ＩＰＴＧ诱导下，测定 了不同克隆株基因表达产物对大肠杆菌细胞生长的影响。结果表明：ＬＺ２０的细胞生长不受影 响；ＬＺｃｙｔＡ的细胞被杀死；ＬＺ２０Ａ的细胞生长也不受影响。这表明２０ｋＤａ蛋白质基因与ｃｙｔＡ 蛋白基因重组后，２０ｋＤａ蛋白质基因表达产物可保护ＣｙｔＡ蛋白对大肠杆菌的溶细胞作用，而 巳这种作用并不因不同大肠杆菌受体而改变。
In order to understand the influence of the 20kDa protein on the cytolytic activity of CytA protein, a set of PCR primers were designed to amplify 20kDa protein and CytA protein genes. The genes were liga0ted to E. colt expression vector pUHE24 and transformed into E coli XLI and DH5. The clones, LZcytA, and LZ20A containing 20kDa protein gene, cytA protein gene and 20kDa-cytA genes were obtained respectivelsy. The influence of the clones on the growth of E. coli scells was determined under induction of IPTG. The results showed that the growth of LZ20 cellss were not affected, LZcytA cells were killed, and the growth of LZ20A cells were not affected. It was suggested that expression product of 20kDa protein gene protected the host cells from the cytolytic effect of CytA protein. This was supported by using different host strains.
关键词
苏云金芽胞杆菌
20kDa蛋白质
cytA蛋白
Bacillus, thuringiensis subsp. israelensis, 20kDa protein gene, cytA protein gene
