Abstract:Abstract: Primary dysmenorrhea can be caused by over-contraction of uterine smooth muscle (USM). Calcium channel, playing crucial roles in USM contraction, is constituted by several key proteins, including voltage-dependent L-type calcium channel subunit beta-1 (CACNB1). To elucidate the roles of miR-16 in regulating USM contraction, it is necessary to know whether rat Cacnb1 gene is a target of miR-16. Firstly, the 3′UTR of rat Cacnb1 was amplified by specifically designed primers and cloned to the dual luciferase reporter vector pmiR-RB-REPORTTM. Meanwhile, the vector with mutant 3′UTR of rat Cacnb1 was also constructed. Then, the constructed vectors were transfected in 293T cells with miR-16 mimics, respectively, followed by the assay of relative luciferase activity. The results demonstrated that miR-16 mimics were able to significantly inhibit the relative luciferase activity by targeting 3′UTR of rat Cacnb1 (n=3, P<0.001). However, no inhibitory activity was shown with the mutation of 3′UTR of rat Cacnb1. It can be concluded that rat Cacnb1 is able to be down-regulated by miR-16. Therefore, rat Cacnb1 is the targeted gene of miR-16.
引用本文:
张小真, 石 科, 李桂玲, 杨 亮, 范 沛. 大鼠Cacnb1报告基因载体构建及miR-16靶向验证[J]. 生命科学研究, 2016, 20(4): 309-313.
ZHANG Xiao-Zhen, SHI Ke, LI Gui-Ling, YANG Liang, FAN Pei. Construction of Reporter Vector for Rat Cacnb1 Gene to Verify Its miR-16 Target. Life Science Research, 2016, 20(4): 309-313.