重组载体_____地中海贫血;_巢式PCR;

人β地中海贫血突变基因重组载体pcDNA3.1LCRβ41/42的构建

http://www.100md.com 《广东药学院学报》 2005年第1期

重组载体,,地中海贫血;,巢式PCR;,β41,42基因;,pcDNA3.1;,重组载体,1材料与方法,2结果,3讨

     摘要目的构建人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5.5 kg β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。

    关键词地中海贫血; 巢式PCR; β41/42基因; pcDNA3.1; 重组载体

    Plasmid construction of the human thalassemic gene β41/42 and LCR gene

    TANG Hui,LI Hongjian,HE Zhenyu, LI Yueqin,TANG Dongsheng,ZHOU Tianhong,CHENG Longqiu

    (Section of Biochemistry and Molecular Biology/Section of Medical Genetics, Jinan University, Guangzhou 510632,China)

    Abstract Objective In order to establish the foundation for in vitro cell and transgenic mouse model, the Human thalassemic gene β41/42 and LCR gene were cloned and sequenced. Methods LCR and the human thalassemic β41/42 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 β41/42 gene in the correct recombinant direction. Sequencing showed that the cloned insert was correct. Conclusion The plasmid pCDNA3.1LCRβ41/42 has been successfully constructed. ......

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