Construction Of Recombinant Plasmids With Different Prostate-specific Membrane Antigen (PSMA) Promoter/enhancer And Choosing Of Transcriptional Activity

Objective: to observe and contrast the transcriptional activity of PSMA promoter/enhancer in different kinds of cell lines, and to choose PSMA promoter/enhancer with the strongest transcriptional activity from different promoter and enhancer combination.Methods: Qamplify PSMA promoter and enhancer DNA sequences fromLNCaP cell by polymerase chain reaction. Then construct recombinant plasmids(pEGFP-PSMA_Pro ,. pEGFP-PSMA_E-p. pEGFP-PSMA_E(r)-p , pEGFP-PSMAE(d)-p and pEGFP-PSMAE(t)-p) with molecular clonal technique.(2) At the same time, all experimental cell lines were analysed for the expression of PSMA using PSMA monoclonal antibody with ABC kit immunohistochemical assay. â‘¢ After plasmids transfected via liposome,observe the expression of reporter gene(EGFP) through fluorescent microscope, compare the different level of EGFP expression with RT-PCR and flow cytometry.Result: â‘  with immunohistochemical stain, only LNCaP cell line expressesPSMA positive. (2)PSMA promoter/enhancer pose transcriptional activity inPSMA(+) cell lines, while without any activity in PSMA(-) cell lines. <3)PSMAe-p poses the strongest activity in different PSMA promoter/enhancercombinations.Conclusion: In our research, we confirm the specific expression of PSMA inprostate cell again. Similarly, transcriptional activity Of PSMApromoter/enhancer pose specificity of prostate, we have chosen the strongestsequence of PSMA promoter/enhancer, which could be utilized in advancedresearch.