利用 TALEN 技术高效制备 TXNIP 基因敲除小鼠模型

摘要

目的：利用显微注射技术注射敲除Txnip基因的TALEN mRNA获得TXNIP敲除小鼠。方法在线设计Txnip敲除位点，构建TALEN载体并在细胞水平验证剪切活性，体外转录TALENs成mRNA并通过显微注射技术注射到C57BL／6J小鼠受精卵，对F0代小鼠进行DNA水平鉴定获得TXNIP敲除小鼠。结果在Txnip第一外显子上设计了TALEN识别剪切位点，并在细胞水平验证具有剪切活性，注射mRNA到受精卵获得了4只敲除小鼠，其中2只Txnip发生移码突变，成功制备了TXNIP敲除小鼠。结论通过注射TALENs mRNA可以高效制备TXNIP敲除小鼠。%Objective To knockout the murine Txnip gene using microinjection of transcription activator-like effector nuclease ( TALEN) mRNAs.Methods TALEN knockout site recognizing Txnip was designed by tools on line, then constructed the vectors and assayed its cleavage activity at cellular level.TALEN mRNA was transcribed in vitro and microinjected into C57BL/6J mouse zygotes.F0 mice were verified at DNA level with BamHI and TXNIP-knockout mice were obtained.Results We designed and constructed TALENs which recognized and cut the first exon of Txnip, and got four TXNIP knockout mice, among which two were frameshift mutation, demonstrating that the TXNIP-knockout mice were generated by TALEN technique.Conclusions Microinjection of in vitro transcribed TALEN mRNAs into murine zygotes is a highly effective and convenient way to develop TXNIP-knockout mouse model.