小鼠Pkd2基因条件性敲除打靶载体的构建

CONSTRUCTION OF TARGETING VECTOR FOR CONDITIONAL KNOCKOUT OF MURINE PKD2

[目的]构建小鼠Pkd2条件性基因敲除打靶载体,为建立Pkd2基因条件性敲除小鼠模型奠定基础。[方法]以正常小鼠(129x1/SvJ)基因组DNA为模板,扩增小鼠Pkd2基因一段包括第9号外显子的长为9.3kb的序列,通过引入LoxP和Neo基因等步骤,建立条件性敲除Pkd2第9号外显子的条件性基因打靶载体。[结果]经多个限制性核酸酶酶切鉴定和测序证实,所构建的Pkd2基因条件性敲除打靶载体符合设计要求。[结论]成功构建小鼠条件性Pkd2基因敲除打靶载体,为建立Pkd2基因条件性敲除小鼠模型奠定基础。

[ Objective] To construct the targeting vector for conditional knockout of polycystic kidney disease 2 gene （Pkd2）. [Methods] A 9.3 kb Pkd2 genomic DNA fragment containing exon 9, obtained from murine genomic DNA（129x1/ SvJ） by PCR, was inserted into T vector （pCR2.1-TOPO） which was used as the homologous ann. LoxP and Neo were inserted for conditional knockout of exon 9 of Pkd2. （Resutts ] The correct structure of the targeting vector was confirmed by restriction enzyme digestion and sequencing analysis. [ Conclusion ] A targeting vector for conditional knockout of routine Pkd2 has been successfully constructed. The construction of targeting vector paves the way for conditional knockout mouse strain generated by targeted mutation of Pkd2.