猪α1,3-半乳糖转移酶基因定向缺失质粒的构建  

Construction of porcine alpha 1,3-galactosyltransferase gene knockout vector

作　　者：常宏宇[1] 宫锋[1] 季守平[1] 王颖丽[1] 郑长青[1] 田曙光[1] 章扬培[1] 

机构地区：[1]军事医学科学院野战输血研究所,北京100850

出　　处：《军事医学科学院院刊》2007年第5期436-439,455,共5页Bulletin of the Academy of Military Medical Sciences

基　　金：国家重点基础研究发展规划资助项目("973"计划)(No.2002CB713804)

摘　　要：目的:构建猪α1,3-半乳糖转移酶(α1,3-galactosyltranferase,GGTA1/α1,3-GT)基因的定向缺失质粒,用于GGTA1基因敲除。方法:以原代猪胚胎成纤维细胞(porcine fetal fibroblast,PFFb)基因组DNA为模板,采用长程PCR方法扩增出GGTA1基因同源长、短臂,将长臂段在NotⅠ位点插入PBSLoxPNEOLoxP质粒中PGK-NEO-PolyA序列的5′端,将短臂段在SalⅠ和XhoⅠ位点插入该质粒PGK-NEO-PolyA序列的3′端。结果和结论:成功构建了猪GGTA1基因的定向缺失质粒NEOSL/GT,为下一步获得GGTA1基因敲除猪打下了基础。Objective： To construct the pig α1, 3-galactosyltransferase （GGTA1/αl, 3-GT） gene knockout vector. Methods ：Two GGTA1 genomic fragments were generated by PCR from purified genomic DNA of primary porcine fetal fjbroblasts（PFFb）. The long fragment was inserted into the Not I site at 5＇ end of the PGK-NEO-PolyA sequence in PBSLoxPNEOLoxP plasmid, and the short fragment was inserted into the Sal I -Xho I site at 3＇ end of the PGK-NEO-PolyA sequence in the plasmid. Results and Conclusion：A GGTA1 knockout vector NEOSL/GT was constructed. This laid a foundation for targeting knockout of the GGTA 1 gene in pig.

关 键 词：移植 异种 α1 3-半乳糖转移酶基因 基因打靶 聚合酶链反应 

分 类 号：Q78[生物学—分子生物学]