Abstract: tTo investigate the effects of miRNAs on the post-transcriptional regulation of circadian genes, we first constructed the clock and bmal1 3′UTR recombinant plasmid. The 3′UTR sequence of mouse clock and bmal1 genes searched from the NCBI was awplified, and then cloned into the pGL3-promoter plasmid. Plasmid DNA was subsequently sequenced to ensure the authenticity and direction of the inserted clock and bmal1 3′UTR. Next, the miRNAs modulation clock and bmal1 expression was analyzed by detecting the luciferase activity. The miRNAs targeting clock, bmal1 gene 3′UTR was predicted by TargetScan. The luciferase activity was analyzed in 293T cells which were cotransfection with recombinant plasmid and miRNA mimic by LP2000. Result showed that the sequences analyzed by RVP4 primer were inversely complementary to the sequences in NCBI. The mouse clock and bmal1 3′UTR recombinant plasmids were successfully constructed, which may lay a foundation for further study of post-transcriptional regulation mechanism of circadian genes. Also the results showed that the luciferase activity of clock and bmal1 3′UTR recombinant plasmid was reduced 33% and 45% by miR-199a-5p（P<0.05）and miR-142-3p(P<0.01)， respectively. The activity of clock and bmal1 gene was modulated by miR-199a-5p and miR-142-3p, respectively.

Key words: clock gene, bmal1 gene, dual luciferase reporter, miR-199a-5p, miR-142-3p