新的白血病相关基因LRP15真核表达载体构建及在K562细胞中的表达

 目的构建LRP15基因开放阅读框架(ORF)序列的真核表达载体,并观察其在K562细胞中的表达情况.方法采用RT-PCR方法自1例白血病患者中克隆LRP15cDNA片段,将目的基因LRP15克隆入pGEM-T载体中并进行酶切鉴定和DNA序列测定.将鉴定过的DNA片段插入真核质粒pcDNA3.1(+),构建真核表达载体pcDNA-LRP15.脂质体介导法将其转染K562细胞,72 h以RT-PCR方法检测转染细胞中LRP15基因的表达情况.结果酶切鉴定和序列分析证实,重组质粒含有人LRP15基因ORF序列;转染实验表明LR15基因能在K562细胞中表达.结论成功构建LRP15基因真核表达载体,并在K562细胞中获得表达,为进一步研究LRP15基因的功能奠定了基础.

Construction of eukaryotic expression vectors of human LRP15 gene and its expression in K562 cells

Objective To construct an eukaryotic expression vector containing the open reading frame (ORF) of LRP15 gene and to detect its expression in K562 cells.Methods The LRP15 gene was ligated with the cloning vector pGEM-T; then, the indentified gene was sub-cloned into eukaryotic expression vector pcDNA3. 1( + ). K562 cells were transfected with the recombinant pcDNA3. l( + )-LRP15 by super-fect(r). The LRP15 expression in the transfected K562 cells were assayed by RT-PCR. Results Restriction analysis and DNA sequencing showed that the recombinant plasmid contained the ORF of LRP15 gene. Transfection experiment verified that LRP15 gene could be expressed in K562 cells. Conclusions The eukaryotic expression vector containing the LRP15 gene was successfully constructed and expressed, which is a basis for the further study of the LRP15 gene function.