Establishment of prion deficient cells in cattle

动物克隆技术已经使几个物种的基因中断成为可能。在这里，我们报告的第一个成功破坏朊病毒基因的牛。方法：利用日本黑牛（JBC）朊蛋白基因构建PrP-5。1和PrP-5。2，牛朊病毒基因的靶向载体，并通过脂质体转染法将这些构建体导入胎儿成纤维细胞系JBC 906中。从833个G418抗性菌落中获得了10个聚合酶链反应（PCR）阳性菌落，Southern印迹分析显示这些菌落中朊病毒基因座成功同源重组。结论：在JBC中进行了朊病毒基因的杂合敲除，目前正在进行通过第二轮靶向906 htGT产生纯合朊病毒敲除JBC。

Animal cloning techniques have enabled gene disruption in several species. Here, we report the first successful disruption of the prion gene in cattle. METHODS : The prion gene of the Japanese Black cow (JBC) was used to construct PrP-5. 1 and PrP-5. 2, the targeting vectors for the bovine prion gene, and these constructs were introduced into the fetal fibroblast cell line JBC906 by the lipofection method. Ten polymerase chain reaction (PCR)-positive colonies were obtained from 833 G418-resistant colonies, and Southern blot analysis revealed successful homologous recombination at the prion locus in these colonies. CONCLUSIONS : Heterozygous knockout of the prion gene was performed in JBC, and production of a homozygous prion knockout JBC by a second round of targeting 906htGT is currently in progress.