关键词: CYP2J3, 载体构建, 心肌细胞, 转染

Abstract: Objective To clone rat CYP2J3 gene and recombine eukaryotic expression vector, pcDNA3.1(+)-CYP 2J3, transfect the vector by FuGENE HD transfection agent, and measure its expression level in myocardial cells of rats. Methods The total RNA was extracted from rat liver. The cDNA of CYP2J3 was amplified by RT-PCR and OE-PCR, and inserted into pBS-T vector. The recombinants were checked by PCR and digestion of restriction endonuclease. The CYP2J3 gene was confirmed by DNA sequencing and cloned into eukaryotic expression vector of the pcDNA 3. 1(+). Resulting in pcDNA3.1(+)-CYP 2J3. Cultured myocardial cells of rats were transfected with pcDNA 3. 1(+)-CYP2J3 by FuGENE HD transfection agent. RT-PCR was used to detect the expression of pcDNA 3. 1(+)-CYP2J3. Results CYP2J3 gene was obtained by RT-PCR and OE-PCR. The recombinant eukaryotic expression vector for CYP2J3 gene had been successfully constructed. The transfected myocardial cell of rats could express CYP2J3. Conclusion Successful cloning of CYP2J3 gene construction of pcDNA3. 1(+)-CYP2J3, and lasting expression in cultured myocardial cells of rats at 24 h, 48 h and 72h after transfecting pcDNA3. 1(+)-CYP2J3 may provided the foundation for the further study of CYP2J3 gene, especially its function study in cells.

Key words: CYP2J3, vector construction, myocardial cell, transfection