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中文摘要：
目的 构建JTV1基因真核表达质粒并稳定转染人白血病细胞系K562,检测转染细胞中JTV1基因mRNA和蛋白的表达水平及其对K562细胞增殖的影响。方法从人外周血单个核细胞中克隆JTV1基因,并将其插入pcDNA3.1表达载体中,构建真核重组表达质粒pcDNA3.1-JTV1,经脂质体介导转染K562细胞,采用RT-PCR和Western blot法鉴定转染细胞中JTV1基因mRNA和蛋白的表达水平;MTT法检测JTV1稳定表达对K562细胞增殖的影响。结果重组表达质粒pcDNA3.1-JTV1经双酶切及测序证实,目的基因已插入质粒中;人JTV1基因能在K562细胞中稳定表达;JTV1具有抑制K562细胞增殖的作用。结论已成功构建了JTV1基因真核表达质粒,并获得了稳定表达人JTV1基因的K562细胞克隆,为进一步研究人JTV1基因的功能及其与白血病细胞增殖及凋亡的相关性提供了细胞模型。
英文摘要：
Objective To construct a eukaryotic expression vector for JTV1 gene,stably transfect to human leukemia cellline K562,determine the expression levels of JTV1 mRNA and protein and investigate the effect of expressed product on proliferationof K562 cells. Methods JTV1 gene was cloned from human peripheral blood mononuclear cells（PBMCs）and inserted into expression vector pcDNA3. 1. The constructed plasmid pcDNA3. 1-JTV1 was transfected to K562 cells in mediation of liposome. The ex-pression levels of JTV1 mRNA and protein were determined by RT-PCR and Western blot respectively. The effect of stable expressionof JTV1 on proliferation of K562 cells was determined by MTT method. Results Restriction analysis and sequencing proved that target gene was inserted into recombinant plasmid pcDNA3. 1-JTV1. JTV1 was stably expressed in K562 cells,which inhibited the pro-liferation of K562 cells. Conclusion The eukaryotic expression vector for JTV1 gene was successfully constructed,and the K562cell clones stably expressing JTV1 gene was obtained,which provided a cell model for further study on the function of human JTV1gene as well as its relationship to the proliferation and apoptosis of leukemia cells.
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