四环素抗性重组鼠李糖乳杆菌的构建

Construction of Recombinant Lacbacillus rhamnosus with Tetracycline Resistance

将含有鼠李糖乳杆菌同源序列的自杀质粒pUC-ldhD-Ter转化到鼠李糖乳杆菌JCM 1553中,成功获得2株含四环素抗性的重组鼠李糖乳杆菌GL-1和GL-2。采用PCR技术鉴定重组菌株染色体基因组上含有四环素抗性基因Ter;生长曲线说明四环素抗性基因的插入对鼠李糖乳杆菌的生长没有较大影响。抗性菌株GL-1和GL-2在含10%葡萄糖的摇瓶发酵培养基发酵结果显示:37℃,200 r/min发酵40h,菌体密度OD600最大达到25.49和24.66,残糖含量为0.60%和0.63%,L-乳酸最高产量为93.956 g/L、93.693 g/L,葡萄糖转化率达到94.82%、94.56%,与原始菌株没有显著区别。

The suicide plasmid pUC-ldhD-Ter containing homologous sequence from Lacbacillus rhamnosus was transformed into Lactobacillus rhamnosus JCM 1553,Two recombinant stains have been constructed successfully and named GL-1,GL-2.PCR was applied to clone the gene of tetracycline from the chromosomal DNA of recombinant stains.The growth curve of the recombinant Lacbacillus rhamnosus was consistent with that of the wild-type strain.The highest yield of L-lactic acid in Erlenmeyer flasks from 10% glucose at 37℃/200 r/min by GL-1 and GL-2 are 93.956 g/L,and 93.693 g/L after 40h.The conversion of glucose are 94.82% and 94.56%.OD600 value of the two strains achieve 25.49 and 24.66.The residual sugar content are 0.60% and 0.63% respectively.The result showed no remarkable difference from the wild type.