摘要
根据GenBank报道的人源温和气单胞菌溶血素(hly)基因序列设计引物,经PCR扩增,得到大小约为1470 bp的阳性产物.利用BamHⅠ,HindⅢ位点将hly基因片段克隆到pMD19-T载体,构建得到重组质粒pWT-hly,并比较所克隆基因序列的同源性.然后将该基因克隆到原核表达载体pET-28a,构建原核重组表达质粒pWET-hly,转化BL21(DE3)并进行SDS-PAGE分析.结果表明:克隆的hly基因与GenBank报道的溶血素基因同源性为96.19%;重组菌株经IPTG诱导后,hly基因得到了高效表达.
According to the published hemolysin nucleotide sequence from human Aeromonas sobira in GenBank, a pair of primers were designed and synthesized, which were localized in conservative regions of hemolysin gene and were modified with Barn H Ⅰ and Hind Ⅲ enzyme sizes. A 1 470 bp gene fragment was amplified via PCR and the recombinant plasmid pWT-hly was constructed. Nucleotide sequence analysis indicated that the gene was 1 467 bp in length and homology of nucleic acids between this gene and other similar gene in GenBank was 96.19 %. The recombinant expression plasmid pWET-hly was constructed and transformed into E. coli BL21 （DE3）. Then the recombinant strain BL21 （DE3） （pWET-hly） was obtained and SDS-PAGE analysis indicated that the hly gene was highly expressed when the recombinant strains was induced by IPTG.
出处
《河北师范大学学报（自然科学版）》
CAS
北大核心
2008年第2期239-242,共4页
Journal of Hebei Normal University：Natural Science
基金
河北省自然科学基金(C2007000263)
