人β地中海贫血突变基因重组载体pcDNA3.1-LCR-β^(41/42)的构建

Plasmid construction of the human thalassemic gene β^(41/42) and LCR gene

目的构建人β地中海贫血突变基因β41 /42及具有调控功能的基因座控制区(LCR)重组载体,为该病基因治疗的体外细胞模型和转基因小鼠模型的建立奠定基础。方法抽提β41 /42纯合子患者的DNA,PCR扩增人β珠蛋白基因座控制区 (LCR)和β41 /42基因,将其串联克隆至pcDNA3. 1 中,酶切及测序鉴定重组载体。结果所构建的重组载体中含人β珠蛋白基因座控制区(LCR)和β41 /42基因,测序结果及引入方向正确。结论成功构建了含 5. 5kgβ珠蛋白基因座控制区(LCR)的β41 /42人重型地中海贫血基因的重组载体。

Objective In order to establish the foundation for in vitro cell and transgenic mouse model, the Human thalassemic gene β 41/42 and LCR gene were cloned and sequenced. Methods LCR and the human thalassemic β 41/42 Gene were amplified by PCR, and cloned into the plasmid pcDNA3.1. The recombinant plasmid was certified by enzyme digestion and sequencing. Results A recombinant plasmid was obtained, which contained LCR and the human thalassemic β 41/42 gene in the correct recombinant direction. Sequencing showed that the cloned insert was correct. Conclusion The plasmid pCDNA3.1-LCR-β 41/42 has been successfully constructed.