Journal: BMC Cancer
Article Title: Functional promoter upstream p53 regulatory sequence of IGFBP3 that is silenced by tumor specific methylation
Figure Lengend Snippet: Methylation analysis of IGFBP-3 promoter for induction by p53 . A, Schematic of four wild type p53 consensus sequence of IGFBP-3 (-159/-209) of pGL2-210 and a sequence of the methylated construct carrying the CpG methylation in the core consensus sequence (underlined) of pGL2-210. B, HepG2 cells were transiently transfected with wild type IGFBP-3 promoter-luciferase reporter construct or methylated IGFBP-3 promoter-luciferase reporter construct with (filled box) or without (open box) co-transfection by pCMV-p53, and luciferase activity was measured after 48 h. Mean count of luciferase activity +/- SEM is shown.
Article Snippet: A series of deletion mutant constructs, pGL2-270, pGL2-240, pGL2-210, pGL2-180, pGL2-150, pGL2-120, pGL2-90, pGL2-60, pGL2-30, and pGL2-1 containing the indicated fragments upstream of the transcription start site and 60 bp of fragments downstream of the transcription start site, were generated by PCR amplification of the promoter fragment and subsequent subcloning of the Mlu I-Bgl II fragment to pGL2-Basic (Table , Fig. ).
Techniques: Methylation, Sequencing, Construct, CpG Methylation Assay, Transfection, Luciferase, Cotransfection, Activity Assay