摘要
为探讨lncMD促进牛成肌细胞分化的分子机制,利用UCSC基因组浏览器分析lncMD的基因组结构,通过qPCR技术检测lncMD在牛不同组织中的表达量,采用过表达、干扰技术研究牛成肌细胞中lncMD对miR-1表达的调控作用,并通过双荧光素酶报告基因试验研究miR-1与靶基因Pax7的靶向关系。结果发现,lncMD的第三外显子可以编码miR-1,且两者均在牛的骨骼肌中高表达,lncMD正调控miR-1表达;生物信息学分析发现,Pax7(paired box 7)3'UTR区有4个miR-1的结合位点;双荧光素报告系统结合定点突变试验证实,牛Pax7基因是miR-1的下游靶基因。综上,本研究阐明lncRNA lncMD通过编码miR-1抑制其靶基因Pax7的功能,进而促进成肌细胞分化,这些发现可为肉牛的遗传改良和精准分子设计育种奠定科学基础。
The purpose of this study was to explore the molecular mechanism by which lncMD promotes bovine myoblast differentiation.The UCSC genome browser was utilized to analyze the genomic structure of lncMD.Quantitative real-time PCR was employed to detect the expression levels of lncMD in different bovine tissues.By utilizing the overexpression and interference strategies,the regulatory effect of lncMD on miR-1 expression was analyzed in primary cultured bovine myoblasts.The targeting relationship between miR-1 and its target gene Pax7 was further detected through dual luciferase reporter gene assay.The results showed that the third exon of lncMD can encode miR-1,both of which are highly expressed in bovine skeletal muscle.lncMD can positively regulate miR-1 expression.Bioinformatics analysis indicated that there are four miR-1 binding sites in the 3'UTR region of Pax7(paired box 7)gene.The dual-fluorescence assay combined with site-specific mutation testing confirmed that the bovine Pax7 gene is a downstream target gene of miR-1.Based on the above results,this study demonstrated that lncRNA lncMD can encode miR-1 and inhibit the function of its target gene Pax7,thereby promoting myoblast differentiation.The results can lay a foundation for genetic improvement and precise molecular design breeding in beef cattle.
作者
孙晓梅
王奕猛
徐杰
毛秋彤
李明勋
蓝贤勇
SUN Xiaomei;WANG Yimeng;XU Jie;MAO Qiutong;LI Mingxun;LAN Xianyong(College of Animal Science and Technology,Yangzhou University,Yangzhou,Jiangsu 225000;College of Animal Science and Technology,Northwest A&F University,Yangling,Shaanxi 712100,China)
出处
《家畜生态学报》
北大核心
2023年第12期15-20,共6页
Journal of Domestic Animal Ecology
基金
国家自然科学基金(No.32202625,No.32372852)
山东省农业良种工程项目(No.2020LZGC014)。
