人CD160基因转染细胞株的建立及其生物学特性的研究

Establishment of Transgenic Cell Lines of human 160 gene and Preliminary Study of Its Biological Characteristics

目的克隆人CD160基因,并构建含有该目的基因的pIRES2-EGFP重组质粒载体,获得稳定表达CD160分子的基因转染细胞。方法从人外周血cDNA文库中扩增CD160的基因,另以VSIG4基因的cDNA为模板扩增跨膜区基因片段,将它们一并装入pIRES2-EGFP质粒载体中,脂质体法共转染L929细胞,用G418筛选出能稳定表达CD160分子的L929细胞株。结果成功克隆出了CD160的基因,构建了pIRES2-EGFP/CD160TMV质粒载体,并建立了稳定表达人CD160分子的L929转基因细胞株,该转基因细胞能结合L929/HVEM转基因细胞,证明其功能性。结论构建了含人CD160基因的重组pIRES2-EGFP质粒载体和建立稳定表达人CD160分子的细胞株,为CD160分子的后续研究奠定基础。

Objective To clone human CD160 gene and construct recombinant plasmid vector carrying the target gene which could be expressed stably in cell line L929.Methods The CD160 gene was amplified by RT-PCR from human peripheral blood mononuclear cells.Besides,transmembrane region gene was amplified by PCR using the template from the whole cDNA of VSIG4 gene.And it was inserted into eukaryotic expressing vector pIRES2-EGFP.The recombinant vector was then transfected into L929 cells.After being selected by G418,transgenic cell line expressing stably human CD160 was established.Results The gene of CD160 was successfully cloned,and its recominant vector pIRES2-EGFP/CD160TMV was constructed.The transfectant stably expressing human CD160 protein on the cells membrane was established successfully.This transfectant could bind to another transgenic cell line L929/HVEM through co-culture.Conclusion The recombinant vector and the transgenic cell lines stably expressing human CD160 molecule on the cells surface has been obtained and can be used for further biological function research.