利用Red同源重组系统进行牛β酪蛋白基因敲除.pdf

HEREDITAS (Beijing) 2007 5 , 29(5): 570―574 ISSN 0253-9772 研究报告 DOI: 10.1360/yc-007-0570 Red 1,2 3 1 2,4 2 薛可 , 李峰 , 罗光彬 , 黄玮玮 , 陈学进 1. , 110161; 2. , 200092; 3. , 200031; 4. , 510642 : 利用 EL350 基因工程菌进行同源重组, 成功进行基因敲除已有报道, 但利用该系统进行乳腺生物反应器 质粒构建的研究却未见报道。实验采用含有完整的牛β 酪蛋白基因的 CSN2 质粒作为基因打靶的载体, 设计不 同的同源臂, 成功地敲除了β 酪蛋白基因的编码区。并且同时利用同源重组技术对敲除不同大小的 DNA 片段 的效率进行了研究。为进一步利用 CSN2 质粒两端的调控序列, 插入新的基因, 研究其表达功能, 或者进行乳 腺生物反应器的研究奠定了基础。 : 同源重组; 牛β酪蛋白基因; Cre-loxP 重组系统; 基因敲除 Knock out of bovine beta casein gene by homologous recombination 1,2 3 1 2,4 2 XUE Ke , LI Feng , LUO Guang-Bin , HUANG Wei-Wei , CHEN Xue-Jin 1. College of Animal Science and Vet Medicine of Shenyang Agricultural University, Shenyang 110161, China; 2. Shanghai Jiao Tong University Xinhua Hospital Development Biology Research Center , Shanghai 200092, China; 3. Institutes of Biochemistry and Cell Biology, SIBS, CAS, Shanghai 200031,China; 4. College of Animal Science of Huanan Agricultural University, Guangzhou 510642, China Abstract: It has been reported that homologous recombination with Red system has been successfully used for knock-out. We try to work on the construction of the expression vector of Mammary Gland with Red system. This study takes CSN2 as a vector for gene target, which contains the complete bovine beta casein gene. Different homologous arms were designed and the CDS region of the beta casein gene was successfully knocked out. The efficiency was also explored for knocking out different DNA fragment. Based on the study, it is very conve