重组pQE30 tPAr质粒在JM109菌株中的稳定性研究

重组pQE30 tPAr质粒在JM109菌株中的稳定性研究 Study on the Heredity Stability of a Recombinant Plasmid pQE30tPAr in E.coli JM109 贺石汉 HE Shi-han 1 1 2 3 4 first-author 李宝宗 LI Bao-zong 1 1 2 3 4 郜金荣 GAO Jin-rong 1 1 2 3 4 叶林柏 YE Lin-bai 1 1 2 3 4 郑永勋 ZHENG Yong-xun 2 1 2 3 4 佘应龙 SHE Ying-long 1 1 2 3 4 吴正辉 WU Zheng-hui 1 1 2 3 4 武汉大学生命科学学院病毒研究所 武汉大学生命科学学院病毒研究所 Institute of Virology,College of Life Sciences,Wuhan University Institute of Virology,College of Life Sciences,Wuhan University 理科大学生物系 理科大学生物系 Biology Department,Nature Science University,Pyongyang,The Democratic People’s Republic of Korea Biology Department,Nature Science University,Pyongyang,The Democratic People’s Republic of Korea 武汉大学生命科学学院病毒研究所,湖北,武汉,430072 武汉大学生命科学学院病毒研究所,湖北,武汉,430072 ,平壤,朝鲜 ,平壤,朝鲜 将构建好的含有人变异tPA基因的重组pQE30tPAr质粒的JM10 9菌株原始种子库菌种 ,在含Amp的平板培养基划线法连续传代至 10 0代 ,其在固体培养基中生长速度、菌落形态和菌体形态、Amp抗性等方面与原始种子库无明显差异 ;每隔 10代提取质粒DNA用HindⅢ和KpnⅠ限制性内切酶切割检查 ,酶切图谱没有改变 ;DNA测序未见tPAr基因变异 .用原代和第 10 0代培养诱导表达tPAr,表达水平和tPAr活性均无明显差异 .菌体的SDS pAGE图谱也完全相同 .以上结果表明所建立的JM 10 9tPAr工程菌株具有良好的遗传稳定性 The strain of JM109pQEtPAr from the original stock which harbering a recombinant plasmid pQEtPAr including mutation in tPA gene had been subcultured serialy to 100 th generation in Amp.plate. The growth characteristics and the morphology of the cultures did not show differences among the generations.Every 10 generation the digestion products of pQEtPAr plasmid by HindIII and Kpn1 restriction enzymes are same. as well as DNA sequenes of the tPA gene shows no differences among the generations, in addition to that, original strain and 100 th generation strain have a similar tPA expression level where these tPA proteins shows a simillar activity. From the above results,it is indicated that recombinant strain JM109 pQEtPAr has a high hereditable stability. tPAr 工程菌株 传代 遗传稳定性 tPAr engineering strain generations heredity stability R346 国家科技攻关项目 （ 2 0 0 1BA749C）;湖北省重点资助项目 （ 2 0 0 1AA3 0 4B0 1） 2003-04-01 2021-04-01 4