肉牛MSTN基因miRNA干扰载体的构建及干扰效果验证  

Construction of MSTN Gene miRNA Interference Vector and It's Interference Effect Evaluation

作　　者：郑艳玲[1,2] 郑月茂 姜八一[2] 王勇胜 张涌[1] ZHENG Yan-ling;ZHENG Yue-mao;JIANG Ba-yi;WANG Yong-sheng;ZHANG Yong(College of Veterinary Medicine of Northwest A&F University,Yangling,Shaanxi 712100,China;Shandong Vocational Animal Science and Veterinary College,Weifang,Shandong,261061,China)

机构地区：[1]西北农林科技大学动物医学院,陕西杨凌712100 [2]山东畜牧兽医职业学院,山东潍坊261061

出　　处：《动物医学进展》2022年第10期7-11,共5页Progress In Veterinary Medicine

基　　金：转基因生物新品种培育科技重大专项(2008ZX080072004);山东省农业良种工程项目(南种北繁)(2017LZN022)。

摘　　要：构建肉牛MSTN基因的miRNA特异性干扰载体,并在肉牛骨骼肌卫星细胞中验证其有效性。设计4对肉牛MSTN基因特异性miRNA寡聚单链DNA序列,分别克隆在pcDNA^(TM) 6.2-GW/EmGFPmiR上,构建MSTN基因miRNA的干扰载体,转染肉牛骨骼肌卫星细胞,实时荧光定量PCR及Western blot分别用于检测MSTN基因mRNA及蛋白表达量,结果显示,成功构建了4个miRNA干扰载体miR-MSTN-1、miR-MSTN-2、miR-MSTN-3及miR-MSTN-4。4个载体均能有效下调MSTN mRNA及蛋白的表达,其中miR-MSTN-4的干扰效果最好。说明成功构建针对肉牛MSTN基因的miRNA特异性有效干扰载体,为通过MSTN基因的miRNA干扰生产高产肉性能肉牛的研究奠定了基础。To construct specific miRNA interference vectors targeting MSTN gene of beef cattle and detect the suppression efficiency of the constructed vectors in skeletal muscle satellite cells of beef cattle,four pairs of beef cattle MSTN gene specific miRNA oligomeric single-stranded DNA sequences were designed and cloned in pcDNA TM 6.2-GW/EmGFPmiR,the recombinant plasmid was transfected into skeletal muscle satellite cells.The interference efficiency was detected by real-time quantitative PCR and Western blot.The results showed that four interference vectors were successfully constructed,including miR-MSTN-1,miR-MSTN-2,miR-MSTN-3 and miR-MSTN-4.The four vectors can effectively down regulate the expressions of MSTN mRNA and protein,and the interference effect of miR-MSTN-4 is the best.It showed that a specific and effective miRNA interference vector targetting beef cattle MSTN gene was successfully constructed,which lays a foundation for the research on the production of high-yield beef cattle by miRNA interference of MSTN gene.

关 键 词：MSTN基因 MIRNA 干扰载体 干扰效果 

分 类 号：S818.9[农业科学—畜牧学] S858.23[农业科学—畜牧兽医]