绵羊oar-miR-200b对GNAQ基因的表达调控  

Regulation of gene expression of GNAQ by oar-miR-200b in sheep

作　　者：于要升 林杉[1] 王开胜[1] 雷小萍 梁慧慧[1] 杨恒[1] 赵宗胜[1] 

机构地区：[1]石河子大学动物科技学院,新疆石河子832003

出　　处：《石河子大学学报（自然科学版）》2016年第4期424-430,共7页Journal of Shihezi University(Natural Science)

基　　金：国家自然科学基金项目(1100902)

摘　　要：为了探讨mi R-200b是否通过靶向作用GNAQ基因并以此调控发情过程,本研究通过双荧光素酶报告基因系统,验证mi R-200b对靶基因GNAQ的真实干扰作用,并在下丘脑神经细胞和卵巢颗粒的细胞中过表达mi R-200b,用荧光定量PCR检测了GNAQ下游基因的表达变化。结果显示:共转染GNAQ-3'UTR-psi-CHECKTM-2和pri-mi R-200b-pc DNA3.1(+)组的hela细胞荧光素酶值是只转染psi-CHECKTM-2组的77.5%,其余两组与只转染psi CHECKTM-2组之间的差异无统计学意义。通过mi R-200b干扰载体转染下丘脑神经细胞,选择GNAQ下游基因ITPR、PRKCB、GNA1、CREB、MAP3K1、GPR54、KISS1及Gn RH进行实时荧光定量检测,结果发现:mi R-200b干扰组和对照组相比,GNAQ基因显著下调,下游ITPR、PRKCB、GNA1和MAP3K1、发情关键基因GPR54、KISS1和Gn RH显著上调。通过mi R-200b干扰载体转染卵巢颗粒细胞,选择GNAQ下游基因COMT、HSD3B1、CYP1A1和PKA基因进行表达量验证,结果表明:mi R-200b干扰后,GNAQ基因表达量升高,下游基因COMT、HSD3B1、CYP1A1和PKA表达量下降。由此推测,mi R-200b真实靶向调控GNAQ基因,进而调控发情过程。To investigate whether the mi RNA-200 b can target GNAQ genes and regulate the estrous process. The Dual-luciferase vectors was applied to test the interfering effect of mi RNA-200 b on GNAQ genes, and the expression of mi R-200 b gene in hypothalamus nerve cells and ovarian granulosa cells was measured. The GNAQ downstream gene expression was measured by Q-PCR. The results showed that the value of Dual-luciferaseon of the GNAQ-3＇UTR-psi-CHECKTM-2 and pri-mi R-200b-pc DNA3.1（＋） transfection group was 77.5% of that in the psi CHECKTM-2 transfection group. The other two transfected groups were without statistical significance when compared to the psi CHECKTM-2 transfection group. mi R-200 b was used to interfere hypothalamus nerve cells carriers, and real-time Q-PCR was conducted to the downstream genes（ITPR,PRKCB, GNA1, CREB, MAP3K1,GPR54, KISS1 and Gn RH） of GNAQ. Comparing the mi R-200 b interference group, it was found that GNAQ gene significantly decreased, however, ITPR, PRKCB, GNA1, MAP3K1 genes and estrous related gene GPR54 was significantly increased. mi R-200 b was used to interfere hypothalamus nerve cells ovarian granulosa cells, and the downstream genes（COMT, HSD3B1,CYP1A1 and PKA） of GNAQ was measured by Q-PCR. It turned out that GNAQ gene expression was increased, and its downstream COMT, HSD3B1, CYP1A1 and PKA gene expression was decreased. It is concluded that the mi R-200 b may target GNAQ gene and regulate the estrous process by targeting the GNAQ.

关 键 词：绵羊 miR-200b 下丘脑 卵巢 

分 类 号：S813.1[农业科学—畜牧学] Q343[农业科学—畜牧兽医]