用细菌噬菌体T7RNA聚合酶高效表达天蚕蛾菌素(CecropinA)基因

Expression of Cecropin A Genes by Bacteriophage T7 RNA Polymerase

将经聚合酶链反应 (PCR)扩增的 156 bp Cecropin A基因 Ce克隆于载体 p Bluescript(PBS) II KS(+ )上 ,得到重组质粒 p BCe。将该质粒转入 E.coli LE392 .2 3菌株中 ,以含有 T7RNA聚合酶 Gene 1克隆的噬菌体 DE2进行感染表达已克隆的 p BCe基因。提取基因表达产物后进行Western blot分析 ,并进行产物的抗 E.coli JM10 1菌株生长活性试验。结果表明 ,在菌体合成的总蛋白中有 Cecropin A成分存在 ,其含量约占菌体总蛋白的 10 % 。

Cecropin A gene expression system based on bacteriophage T7 RNA polymerase has been developed. 156 bp nucleotides which carring Cecropin A gene produce by PCR were cloned into expression vector pBluescript Ⅱ (PBS)KS(+), Cecropin A recombinant plasmide pBCe was obtained. Then pBCe was transformed into E.coli LE392.23. The transformant was infected by bacteriophage DE2 which contains T7 RNA polymerase from the cloned Gene 1, and then to direct expression of cloned pBCe genes. The expression product were analyzed by western blot and antibacterial activity test by the strain of E. coli JM101. The results showed the presence of Cecropin A in the expression products, and which accounts for about 10% of the total crude protein. This Cecropin A also has antibacterial activity toward the test bacterial strain.