流动引物PCR高效筛选阳性XRCC1基因敲除小鼠细胞的方法

Employing PCR with floating primer to effectively screen XRCC1 gene knock out mice cells

关键词:  基因敲除  流动引物  XRCC1

Key words:  gene knock out  floating primer  XRCC1

摘要:

[目的] 探索流动引物PCR筛选阳性XRCC1基因敲除细胞的可行性。 [方法]构建基因敲除质粒，嘌呤霉素抗性基因5’端加上额外的来自野生型基因被敲除位点内的20 bp DNA序列。使用流动引物（floating primer）和这20 bp序列结合，PCR筛选阳性克隆。[结果]利用流动引物成功筛选小鼠体细胞XRCC1（X射线修复交叉互补蛋白）一个等位基因敲除的阳性克隆子。[结论]流动引物PCR方法可以应用于基因敲除小鼠体细胞的筛选。

Abstract:

[Objective]We attempted to utilize floating primer to screen XRCC1 gene knock out mice cells. [Methods] A gene knock out vector was constructed, in which an additional 20 bp DNA sequence from the knocking-out sites of wild-type gene was added to the 5’ terminus of puromycin resistant gene. A primer named for floating primer was designed to anneal with this 20 bp DNA sequence. [Results]By using floating primer, the mice cells that have one deleted XRCC1 （X-ray repair cross complementing protein1）allele were effectively screened. [Conclusion]PCR with floating primer is a useful method for screening gene knock out mice cells.