Molecular Cloning and Characterization of Human Homeobox Gene Nkx3.1 Promoter

Nkx3.l is a prostate-specific homeobox gene related strongly to prostate development and prostate cancer. To study its regulation of transcription, 1.06 kb 5’ flanking region of Nkx3.1 gene and its 5’ deletion mutants (861, 617, 417 and 238 bp) were obtained by PCR and cloned into pGL3-basic, a promoter-less luciferase reporter vector, to examine their promoter activities driving the reporter gene transcription. pRL-TK, a Renilla luciferase reporter vector was used as internal control, and pGL3-control and pGL3-basic were used as positive and negative control respectively. The promoter activities were determined by dual-luciferase reporter assay 48 h after pGL3 constructs were cotransfected with pRL-TK into prostate cancer cell LNCaP. The results showed that dual-luciferase reporter assay (M1/M2) of pGL3-1.06 kb cotransfection with pRL-TK was 2.7, which was about 1.5-fold higher than that of pGL3-control cotrans-fection with pRL-TK and 50-fold higher than that of pGL3-basic cotransfection with pRL-TK. T