Construction of ERG 3′untranslated region dual-luciferase reporter system and activity dectection
XU Xue-hu
LI Shu-ling
XU Yuan-dong
WU Xiao-bing
WU Shang-biao
CHEN Rong
Abstract：Objective To establish a dual -luciferase reporter vector containing the 3′untranslated region ( 3′UTR) of ERG gene and provide experimental basis for investigation of post -transcriptional regulation of ERG .Methods Genomic DNA was utilized as template for PCR amplification of ERG 3′UTR, which was then inserted into the multiple cloning sites of psiCHECK 2 reporter plasmid .Construction of psiCHECK 2-ERG was confirmed by sequencing .Bioinfor-matic analysis predicted that the 3′UTR of ERG contained a binding site for miR -145-5p.Luciferase activity was detec-ted after cotransfection of miR -145-5p mimics and psi-CHECK2-ERG vector.Cotransfection of mimic control and psiCHECK2-ERG was set as control .Results The dual -luciferase reporter vector containing ERG 3′UTR was con-structed and confirmed by enzyme digestion , electrophoresis and sequencing .Cotransfection of miR-145-5p mimic and dual-luciferase reporter significantly reduced luciferase activity (28.39%, P<0.05).Conclusion The dual-lucifer-ase reporter plasmid containing the 3′UTR of ERG was successfully constructed .Preliminary identification revealed that miR-145-5p regulates ERG at the post -transcriptional level by targeting the 3′UTR of ERG.
Keywords：ERGmicroRNAdual-luciferase reporter vector3′untranslated region
Publication Date：2015-01-01
Online Publishing Date：2025-08-15(First online date of this platform, not the publication date of the document)
Pages：4( 3097-3100 )