Clone and expression of nuclear factor ?B p65 TAD / 第三军医大学学报

ABSTRACT

Objective To acquire NF-?B p65 TAD(transcription activation domain) and construct its eukaryotic expression vector and express it in endothelial cells.Methods Human umbilical vein endothelial cells(HUVECs) were cultured and total RNA was extracted.The p65 TAD gene was amplified by RT-PCR.After sequencing,the p65 TAD gene was inserted into the eukaryotic expression vector pEGFP-N1 with the green fluorescence protein,named pEGFP-N1-p65 TAD.pEGFP-N1-p65 TAD was transfected into HUVECs and its expression was observed under fluorescence microscope and analyzed by Western blotting.Results p65 TAD(288-548) was cloned successfully.The constructed plasmid including p65 TAD gene was identical to the designed.p65 TAD gene was expressed successfully in HUVECs.Conclusion The construction of eukaryotic expression vector including p65 TAD gene and its expression in HUVECs are very successful.