[Construction and expression of recombinant plasmid with prostate-specific membrane antigen promoter and enhancer regulating uracil phosphoribosyltransferase gene].
OBJECTIVE
To construct the recombinant expression plasmid pPSMA(EP)-UPRT including UPRT gene that is regulated by PSMA(enhancer/promoter).


METHODS
By use of PCR, UPRT gene was amplified from E. coli JM109 genome. Then UPRT gene was cloned into the recombinant expression plasmid pPSMA(enhancer/promoter)-EGFP that is driven by prostate-specific membrane antigen promoter and enhancer.


RESULTS
It was found that EGFP was digested by two restriction enzymes from the recombinant expression plasmid pPSMA(enhancer/promoter)-EGFP, and UPRT gene was linked with the recombinant expression plasmid by T4 DNA ligase. We succeeded in constructing the recombinant expression plasmid pPSMA(enhancer/promoter)-UPRT. The recombinant plasmid sequences were verified, and the expression in vitro was measured by MTT.


CONCLUSIONS
The recombinant expression plasmid pPSMA(enhancer/promoter)-UPRT is regulated targetly by prostate-specific membrane antigen promoter and enhancer. It is, of importance to us in studying the UPRT/5-FU for gene therapy of prostate cancer, especially suicide gene therapy (CD/5-FC system).
Zhao
Fu-jun
FJ
Department of Urology, West China Hospital, Sichuan University, Chengdu 610041, China.
Li
Hong
H
Cheng
Hong-ming
HM
Zeng
Hao
H
Wei
Qiang
Q
Li
Xiang
X
D000230:Adenocarcinoma / Q000628:therapy; D000951:Antigens, Neoplasm / Q000096:biosynthesis / Q000235:genetics*; D000954:Antigens, Surface / Q000096:biosynthesis / Q000235:genetics*; D004742:Enhancer Elements, Genetic / Q000235:genetics; D015972:Gene Expression Regulation, Neoplastic; D017930:Genes, Reporter; D043425:Glutamate Carboxypeptidase II / Q000096:biosynthesis / Q000235:genetics*; D049452:Green Fluorescent Proteins / Q000235:genetics; D006801:Humans; D008297:Male; D010430:Pentosyltransferases / Q000235:genetics*; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic; D011467:Prostate / Q000276:immunology* / Q000378:metabolism; D011471:Prostatic Neoplasms / Q000628:therapy; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D014162:Transfection
D000951:Antigens, Neoplasm; D000954:Antigens, Surface; D012333:RNA, Messenger; D011994:Recombinant Proteins; D049452:Green Fluorescent Proteins; D010430:Pentosyltransferases; C023269:uracil phosphoribosyltransferase; C473848:FOLH1 protein, human; D043425:Glutamate Carboxypeptidase II