Identification The Character And Function Of Newly Discovered Negative Regulatory Sequences In Coronary Heart Disease Related SLC22A3 Gene

Objective :To investigate the character and function of two negative regulatory sequences in SLC22A3 gene intron7.Methods: The DNase I sensitivities of the two negative regulatory sequences in the cultured cell lines were analyzed using bioinformatics.Both of the sequences amplified from the bacterial artificial chromosome(BAC)including whole human SLC22A3 gene,were inserted into the interspace between the promoter and enhancer of luciferase reporter gene(LUC)and downstream of enhancer,respectively.Four recombinant plasmids were constructed,and co-transfected with internal reference plasmid pRL-SV40 into HEK293 T cells,with plasmid pGL3-Control used as blank control,then the luciferase activities were detected 24 h after transfection.Results : Both of the two negative regulatory sequences were cis-regulatory elements for being located in the DNaseI hypersensitive sites.Luciferase activities of the four recombinant plasmids were all lowerthan that of pGL3-Control(P<0.05).Conclusions: The results have demonstrated that SLC22A3 gene intron7 contains two negative regulatory elements,that were independent of their position in the gene and acted as silencers,which might provide theoretical basis for further research on the regulation of SLC22A3 gene expression.