重组大肠杆菌生产Taq DNA聚合酶发酵工艺优化 (信息完整）.docxVIP

PAGE PAGE 1 重组大肠杆菌生产Taq DNA聚合酶发酵工艺优化 肖云基金项目：武汉职业技术学院校级项目（2022YK035）作者简介：肖云（1980- 基金项目：武汉职业技术学院校级项目（2022YK035） 作者简介：肖云（1980-），女，副教授，硕士，研究方向为发酵工程。E-mail （1.武汉职业技术学院生物工程学院，湖北 武汉 430072；2.华中农业大学国家微生物重点实验室，湖北 武汉 430070） 摘要：Taq DNA聚合酶是PCR技术中广泛应用的耐热酶。为提高重组大肠杆菌生产Taq DNA聚合酶的产量，本研究以Taq DNA聚合酶产量和酶的比活力为考察指标，通过单因素试验优化产生TaqDNA聚合酶含量的单因素：发酵温度、pH、IPTG诱导剂用量、诱导时间、接种量，结果表明：发酵温度、pH、IPTG诱导剂浓度，对该基因工程菌发酵产Taq DNA聚合酶的比酶活力有显著影响，而诱导培养时间、接种量对产Taq DNA聚合酶的比酶活力影响不显著。故选择发酵温度、pH、IPTG诱导剂浓度作为响应面的三个因素优化重组大肠杆菌生产Taq DNA聚合酶发酵工艺，响应面分析结果显示对酶的比活力显著性顺序为：IPTG诱导剂浓度发酵温度pH，最佳发酵工艺参数为：发酵温度37.2℃，pH为7.5，IPTG诱导剂浓度为0.800mmol/L，在此条件下发酵，Taq DNA聚合酶比活力达到43.822±0.878kU/mg。 关键词：重组大肠杆菌；Taq DNA聚合酶；酶比活力；发酵工艺；响应面分析法 中图分类号：Q815 文献标识码:A Optimization of fermentation process for Taq DNA polymerase production by recombinant Escherichia coli XIAO Yun1,CHEN Jie1, JU Shou-yong1，2 (1. College of Bioengineering, Wuhan Polytechnic, Wuhan,Hubei 430072；2.National Key Laboratory of Microbiology, Huazhong Agricultural University, Wuhan, Hubei 430070) Abstract: Taq DNA polymerase is a thermostable enzyme and widely used in PCR technology. In order to improve the production of Taq DNA polymerase by recombinant Escherichia coli，in this study the enzyme yield and specific activity of the enzyme were taken as the inspection indicators. The single factors producing Taq DNA polymerase content were optimized through single factor test：fermentation temperature, pH, amount of IPTG inducer, induction time, and inoculation amount. The results showed that fermentation temperature, pH, concentration of IPTG inducer had a significant impact on the specific enzyme activity of Taq DNA polymerase produced by the genetic engineering strain, while induction culture time and inoculation amount had no significant impact on the specific enzyme activity of Taq DNA polymerase produced by the genetic engineering strain. So fermentation temperature, pH and IPTG inducer concentration were selected as three factors of response surface to optimize th