钝齿棒杆菌乳酸脱氢酶基因敲除载体构建  

Construction of Corynebacterium crenatum ldhA gene knock-out vector

作　　者：陈小举[1] 李兴江[2] 姜绍通[2] 

机构地区：[1]巢湖学院化学化工与生命科学学院,安徽巢湖238000 [2]合肥工业大学生物与食品工程学院,安徽合肥230009

出　　处：《合肥工业大学学报（自然科学版）》2015年第9期1277-1280,共4页Journal of Hefei University of Technology：Natural Science

基　　金：安徽省高等学校自然科学研究资助项目(Kj2015A216);巢湖学院科研启动基金资助项目(2014003)

摘　　要：乳酸脱氢酶催化丙酮酸生成乳酸是钝齿棒杆菌厌氧发酵产乳酸的关键酶。文章以构建乳酸脱氢酶基因敲除载体为目的,通过在乳酸脱氢酶基因片段中的EcoRV酶切位点插入卡那霉素抗性基因的方法构建钝齿棒杆菌乳酸脱氢酶基因敲除载体。结果显示,成功扩增并克隆到乳酸脱氢酶基因与卡那霉素抗性基因,质粒聚合酶链式反应(polymerase chain reaction,PCR)与酶切结果显示,卡那霉素抗性基因准确插入到乳酸脱氢酶基因中的EcoRV酶切位点,钝齿棒杆菌乳酸脱氢酶基因敲除载体构建成功。When producing lactic acid by Corynebacterium crenatum under anaerobic condition ,the lac‐tate dehydrogenase is the key enzyme which can catalyze the formation of lactic acid from pyruvate .In this paper ,the ldhA gene knock‐out vector was constructed by inserting a km gene into the EcoRV re‐striction site of ldhA .The results indicated that the ldhA and km genes were cloned successfully ,the positive recombinant plasmid was identified by polymerase chain reaction（PCR） and enzyme digestion and the results certified that the km gene was accurately inserted into the EcoRV restriction site of ldhA and the ldhA gene knock‐out vector was successfully constructed .

关 键 词：钝齿棒杆菌 乳酸脱氢酶 基因敲除 载体构建 

分 类 号：Q789[生物学—分子生物学]