|
|||||
|
|
| 可转化人工染色体(TAC)文库载体DNA的制备 | |
| 引用本文： | 李晓玲,李克秀,赵洪锟,赵茂林,董英山.可转化人工染色体(TAC)文库载体DNA的制备[J].生物技术通报,2011(9). |
| 作者姓名： | 李晓玲 李克秀 赵洪锟 赵茂林 董英山 |
| 作者单位： | 1. 吉林省农业科学院生物技术研究中心,长春130033;长春工业大学化学与生命科学学院,长春130012 2. 长春工业大学化学与生命科学学院,长春,130012 3. 吉林省农业科学院生物技术研究中心,长春,130033 4. 中国科学院遗传与发育生物学研究所,北京,100101 |
| 基金项目： | 农业部野生植物资源保护项目(2010); 国家转基因专项(2008ZX08004-004) |
| 摘 要： | 对可转化人工染色体(TAC)pYLTAC747NH/sacB文库载体DNA的制备条件进行了较系统的模索和研究.结果表明,该文库载体经碱裂解法提取、QIAGEN Plasmid Mini kit纯化后,可获得较纯净的载体DNA.对其闭环载体DNA分别用不同酶量的Hinid Ⅲ酶切处理,经琼脂糖凝胶电泳检测得出其最佳Hind Ⅲ完全酶切条件为2 U Hind Ⅲ/μg闭环载体DNA、37℃酶切30 min;分别用0.5 MBU和1 MBU HK脱磷酶/μg对其线性载体DNA进行脱磷处理,经电泳和载体自连产物电转化检测表明其适宜的完全脱磷条件为1 MBU HK脱磷酶/μg线性载体DNA,30℃脱磷1 h；将所制备的线性载体DNA与λ DNA/Hind Ⅲ酶切片段进行连接,连接产物转化频率较高,其电转化大肠杆菌DH10B感受态细胞频率可达到9.6×10s. |
| 关 键 词： | 可转化人工染色体(TAC) 文库载体 酶切 脱磷 |
Preparation of the Vector DNA for Library from the Transformation-competent Artificial Chromosome |
|
| Li Xiaoling,Li Kexiu,Zhao Hongkun,Zhao Maolin,Dong Yingshan.Preparation of the Vector DNA for Library from the Transformation-competent Artificial Chromosome[J].Biotechnology Bulletin,2011(9). | |
| Authors: | Li Xiaoling Li Kexiu Zhao Hongkun Zhao Maolin Dong Yingshan |
| Institution: | Li Xiaoling1,2 Li Kexiu2 Zhao Hongkun1 Zhao Maolin3 Dong Yingshan1(1Biotechnology Research Centre,Jilin Academy Agricultural Sciences,Changchun 130033,2School of Chemistry and Life Science,Changchun University of Technology,Changchun 130012,3Institute of Genetics and Developmental Biology,Chinese Academy of Sciences,Beijing 100101) |
| Abstract: | The optimal condition for the preparation of qualified vector DNA from the transformation-competent artificial chromosome(TAC)pYLTAC747NH/sacB for library was groped and studied.The result showed that the pure vector DNA could be obtained after the plamid was extracted by the alkaline lysis method and purified by QIAGEN Plasmid Mini kit.The closed-loop vector DNA was digested by gradient units of Hind III respectively.The result examined by agarose gel electrophoresis indicated that the optimal digestion co... |
| Keywords: | Transformation-competent artificial chromosome(TAC) Vector for library Digestion Dephosphorylation |
| 本文献已被 CNKI 万方数据 等数据库收录！ | |