Rtn4-A/B基因敲除小鼠模型的制备  

Construction of Rtn4-A/Bknockout mouse model

作　　者：朱莹[1] 胥武剑[1] 宁允叶[1] 吴琳[2] 卢建[3] 李强[1] 

机构地区：[1]第二军医大学长海医院呼吸内科,上海200433 [2]南京军区总医院消化内科,南京200001 [3]第二军医大学基础部病理生理学教研室,上海200433

出　　处：《第二军医大学学报》2013年第8期890-893,共4页Academic Journal of Second Military Medical University

基　　金：国家自然科学基金面上项目(81170060/H011);国家自然科学基金青年项目(81200030/H0108)~~

摘　　要：目的通过制备Rtn4-A/B基因敲除小鼠,探索Rtn4-B基因的生物学功能。方法用细菌人工染色体(BAC)载体构建Rtn4-A/B基因打靶载体并使其线性化,通过电转化法将其转入129SvEv品系雄性小鼠胚胎干细胞(embryonic stem cell,ES细胞)。将正确同源重组的ES细胞注射入C57BL/6J小鼠囊胚腔,繁育出嵌合体小鼠后,进一步繁殖以获得杂合子小鼠。抽提小鼠尾尖组织DNA,采用PCR法鉴定小鼠的基因型。结果基因打靶后,得到14个发生双臂正确同源重组ES细胞克隆。利用阳性ES细胞克隆进行囊胚内显微注射,得到5只嵌合率大于50%的雄鼠,最终繁育得到4只Rtn4-A+/-B+/-杂合子小鼠。结论利用ES细胞基因打靶、同源重组等方法,成功获得Rtn4-A/B基因敲除杂合子小鼠。Objective To generate Rtn4-A/B knockout mouse model and to explore the biological function of the Rtn4 B gene. Methods The targeting construct for inactivating Rtn4-A/B gene was prepared by bacterial artificial chromosome （BAC）. The vector was linearized and electroporated into 129SvEv mouse embryonic stem cells （ES cells）. Then the Rtn4 A/B knockout ES cells were microinjected into blastula of C57BL/6J mice after superovulation. F1 hybrid mice were bred to obtain mouse aggregation chimeras, and were identified by PCR amplification of tail genomic DNA. Results Fourteen clones of gene-targeted ES cells were identified after gene knockout and five male chimeras with a higher than 50 chimeric ratio were produced after microinjection into the blastula. Finally four Rtn4-A/B hybrid mice were obtained. Conclusion A Rtn4-A/B deficient mouse strain has been successfully generated by homologous recombination using genetically modified ES cells.

关 键 词：Rtn4 基因打靶 基因敲除小鼠 胚胎干细胞 同源重组 

分 类 号：R394-33[医药卫生—医学遗传学]