[Construction and identification of recombinant firefly luciferase reporter plasmid pGL2-PEPCK-Luc].
OBJECTIVE
To evaluate in vitro regulation of phosphoenolpyruvate carboxykinase (PEPCK) gene promoter on gene transcription, and construct luciferase reporter plasmid pGL2-PEPCK-Luc.


METHODS
A 550 bp fragment of PEPCK promoter cut from plasmid pPEPCK-int was inserted into transitional vector PBS-SK to construct a transition plasmid PBS-PEPCK. Then the recombinant luciferase reporter plasmid pGL2-PEPCK-Luc was cloned.


RESULTS
Restriction enzymes and nucleotide sequence conformed that the coupling site of recombinant plasmid was correct without base mutation and deletion, and the sequence inserted was the same as data of GeneBank. The luciferase could be expressed in hepatoma cell transfected by pGL2-PEPCK-Luc.


CONCLUSIONS
Established a new means to study transcriptional regulation of PEPCK promoter.
Feng
Kai
K
Department of Endocrinology, PUMC Hospital, CAMS and PUMC, Beijing 100730, China.
Wang
Heng
H
Sun
Qi
Q
Xiao
Xin-hua
XH
D000818:Animals; D001483:Base Sequence; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D006801:Humans; D008113:Liver Neoplasms / Q000235:genetics; D008156:Luciferases / Q000235:genetics*; D008969:Molecular Sequence Data; D010729:Phosphoenolpyruvate Carboxykinase (GTP) / Q000235:genetics* / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011485:Protein Binding; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured
D011993:Recombinant Fusion Proteins; D008156:Luciferases; D010729:Phosphoenolpyruvate Carboxykinase (GTP)