CONSTRUCTION OF EUKARYOTIC EXPRESSION PLASMID OF HEME OXYGENASE-1 GENE AND ITS EXPRESSION IN RAT RENAL TUBULAR EPITHELIAL CELLS

[Objective] To construct HO-1 gene recombinant eukaryotic expression plasmid and to detect the effects of its transfecting into rat renal tubular epithelial cells(RRTECs)and the expression of target protein.Methods Total RNA from rat kidney was extracted.The HO-1 gene was amplified using reverse transcription-polymerase chain reaction(RT-PCR)and cloned into the eukaryotic expression vector pcDNA 3.1(+)by polymerase of recombinant gene technology.Afterwards,the recombinant plasmid pcDNA 3.1(+)-HO-1 was transfected into RRTECs by using the liposome.The effects of transfection and the expressin of HO-1 gene were detected by RT-PCR and Western blot analysis.Results As indentified with double restriction endonuclease digestion and sequence analysis,the DNA sequence of cloned HO-1 gene was correct and the eukaryotic expression recombinant plamid pcDNA 3.1(+)-HO-1 was successfully constructed.RT-PCR and Western bolt analysis showed that HO-1 was transfected successfully into RRTECs and efficiently expressed at both mRNA and protein level.Conclusion The successful construction of eukaryotic expression recombinant plasmid of HO-1 gene and the efficient expression of HO-1 protein in RRTECs lay the foundation for further study on its biological function.