摘要
以酶蛋白表达量、酶活性和比活性为指标 ,从工程菌菌株、转化方法、诱导表达时间、诱导剂浓度 4个方面 ,对TaqDNA聚合酶制备技术进行了优化筛选。用JM 1 0 9菌株制备的总酶活力高于DH5α ,电击法的转化率高于热激法 ,用 1 0mmol/LIPTG诱导 1 2h ,酶蛋白表达量、酶活性和比活性均显著高于其他浓度和时间的处理。SDS PAGE电泳和PCR扩增结果表明 ,用优化技术制备的TaqDNA聚合酶 ,纯度、酶活力和特异性均达到分子生物学实验的要求 。
By using expression product of enzyme protein, enzyme activity and activity ratio as indexes, preparation techniques are optimized through selection of engineering bacterium lines, transformation methods, induction expression time and IPTG concentration. Total enzyme activity prepared with bacterium JM109 is higher than DH5α, transformation ratio of electric shock is higher than heat shock, and enzyme protein expression product, enzyme activity and activity ratio induced by 1.0?mmol/L IPTG for 12?h are significantly higher than other concentration and duration of IPTG induction. The results of SDS-PAGE electrophoresis and PCR amplification indicate that the enzyme product prepared with the optimized techniques meets the requirements of molecular biological experiment in purification, activity and specificity. All the indexes of the product are better than commercial enzyme products in market.
出处
《四川农业大学学报》
CSCD
2004年第4期318-321,共4页
Journal of Sichuan Agricultural University
基金
亚洲玉米生物技术网项目 (AMBIONETICC5 2 4 7B2 4 )
