鱼腥藻PCC 7120染色体上relBE同源基因的克隆及表达

为研究鱼腥藻PCC7120染色体上与relBE同源的基因asl4561和asl4562表达产物的毒素-抗毒素作用,从鱼腥藻细胞中提取总基因组DNA,设计特异引物PCR扩增目的基因asl4561和asl4562,与T载体连接后经XhoI和EcoR I双酶切并与表达载体pET28a（＋）连接,由IPTG诱导表达,并在IPTG浓度和诱导时间上对asl4561基因的表达条件进行了优化.琼脂糖凝胶电泳显示扩增出了264bp的asl4561基因和213bp的asl4562基因,SDS-PAGE检测表明成功表达出15.65kD和13.55kD的两蛋白,当诱导温度28℃、IPTG浓度0.4 mmol/L、诱导时间6h时,asl4561基因表达蛋白在细菌裂解液上清中表达量最大.

Cloning and Expression of the Homologous Gene of relBE in the Chromosome of Anabaena sp.PCC 7120

In order to study the toxin-antitoxin effect of the homologous gene of relBE in the chromosome of Anabaena sp.PCC7120,which are homologous to relBE.Total DNA was extracted from Anabaena sp.PCC7120 cells.The target gene was amplified with PCR,then the genes asl4561 and asl4562 were inserted to pET28a(+) after the double cut-enzymes,the target genes expression was induced by IPTG.Also protein expression conditions were optimized in the concentration of IPTG and induction time.The result of agarose gel electrophoresis showed that asl4561 and asl4562 were cloned and SDS-PAGE indicated two proteins of 15.65kD and 13.55kD were expressed successfully.High yield of asl4561 gene expression protein was achieved by inducing 6 h with 0.4 mmol/Lol/L IPTG at 28℃.