Construction of an expression vector directed by human U_6 small nuclear RNA promoter and identification of expression in gastric carcinoma cell / 西安交通大学学报(医学版)

ABSTRACT

Objective To construct an expression vector directed by hU_(6)snRNA promoter for synthesizing small RNA and to identify its functional activity in the gastric carcinoma cells——SGC-7901.Methods Using human genomic DNA as template,U_(6) snRNA promoter was obtained by PCR method,and then cloned into PUC19 vector to produce the recombinant plasmid PUC-hU_6-extra,which was sequenced and then transfected into gastric carcinoma cell——SGC-7901 with liposome.The effect of expression directed by U_6 promoter was detected by RT-PCR method,and the cell proliferation curve analysis was performed by stained dye.Results The hU_6 snRNA promoter with the first 27 nucleotides followed were successfully cloned into PUC19 plasmid.The recombinant vector could efficiently transcribe small RNA molecules and exerted no effect on cell proliferation in SGC-7901 cells in vitro.Conclusion We have successfully constructed the recombinant PUC-hU_6-extra plasmid vector that can efficiently transcribe small RNA molecules directed by hU_6 snRNA promoter in the gastric carcinoma cells——SGC-7901.