Ⅱ型猪链球菌体内诱导基因表达文库的构建

Construction of a Fusion Gene Expression Library in Screening in Vivo-induced Genes in Streptococcus suis serotype 2 from Swine

为构建用于筛选Ⅱ型猪链球菌体内诱导基因的融合基因表达文库。采用自杀载体pGP704为骨架，将无启动子的cat基因作为报告基因克隆到自杀载体pGP704上，构建报告质粒pGPT04-cat将Ⅱ型猪链球菌基因组DNA的500～1000bp酶切片段克隆到报告质粒pGP704-cat中cat基因的上游，转化受体菌获得融合基因文库；将文库质粒电转化Ⅱ型猪链球菌，得到融合基因表达文库。结果表明：构建的融合基因表达文库以氯霉素作为选择标记，经体外培养，获得2％的菌株具有氯霉素抗性，融合基因表达文库存在可以启动cat基因表达的启动子序列，cat基因在体外可以表达。构建的融合基因表达文库可以用于Ⅱ型猪链球菌体内诱导基因的筛选。

Aim to identify in vivo-induced genes in Streptococcus suis serotype 2 （SS2） from swine, a fusion gene expression library was constructed. Based on the suicide vector pGP704, pGPT04-cat was constructed with promoterless cat gene as reporter gene. Then random DNA fragments （500-1000 bp）, which obtained from SSz genomic DNA partially digested, using Sau3A Ⅰ enzyme, were subcloned into the Bgl Ⅱ site of pGP704-cat. After transformed into the receptor bacterium E. coli, the fusion gene library with SS2 fragments was constructed. Upon introduction of plasmids from the fusion gene library into SS2 strain by electro transformation, we achieved a fusion gene expression library. Cultured the fusion gene expression library on THB medium with chloramphenieol as select marker, the percentage of Cm strains was 2 %. The results indicated that the DNA fragments of SS2 upstream of the eat reporter gene successfuUy got a promoter and the cat gene expressed correctly in vitro. The fusion gene expression library could be used for screening in vivo-indueed genes of SS2 from swine.