摘要
HSL基因所编码的激素敏感性脂酶对脂质代谢具有重要的调控作用,不仅能催化水解脂肪组织中的甘油三酯,还能水解不同组织的胆固醇酯,在机体能量供应和类固醇生成中发挥重要作用。为进一步研究HSL基因功能,本试验设计并构建4个HSL基因shRNA干扰载体,并将干扰载体转染到牛胎儿成纤维细胞中以及和设计构建过表达载体共转染到293T细胞中,转染48h后,分别检测细胞内牛HSL基因的mRNA和蛋白质表达水平,筛选出具有较高干扰效率的shRNA表达载体。结果表明,在牛胎儿成纤维细胞中,4个shRNA载体在转染48h后,shRNA-731组细胞中HSL基因的mRNA表达量显著低于shRNA NC对照组(P<0.05),且干扰效率最佳;同时,在shRNA载体与过表达载体共转染293T细胞中,HSL基因的mRNA和蛋白表达水平显著低于shRNA NC对照组(P<0.05)。本研究为进一步探讨HSL基因功能提供了有效工具,也为研究其与牛脂肪代谢相关关系奠定了理论基础。
The hormone-sensitive lipase encoded by HSL gene plays an important role in lipid metabolism regulation.It can not only catalyze the hydrolysis of triglycerides in adipose tissue,but also hydrolyze cholesterol esters in different tissues,which could provide energy supply as well as steroid production in the body.In order to further study the function of HSL,four HSLgene shRNA interference vectors and overexpression vectors were designed and transfected by transient transfection in bovine fetal fibroblasts as well as into 293T.After transfection for 48h,bovine HSL mRNA and the expression level of protein in cell was screened for shRNA expression vector with high interference efficiency.These results showed that in bovine fetal fibroblasts,mRNA expression of HSL gene in shRNA-731group was significantly lower than that of shRNA NC control group(P<0.05)after 48htransfection,and the interference efficiency was the best.Meanwhile,in the 293Tcells co-transfected with shRNA vector and overexpression vector,the mRNA and protein expression levels of HSLgene were significantly lower than that of shRNA NC control group(P<0.05).This study provides an effective tool for the functional validity of HSLgene in further,and lay a foundation for study of bovine fat metabolism as well as the mechanism of HSL gene.
作者
靳子康
房希碧
高振
潘子意
刘娟
姜平
杨润军
赵志辉
JIN Zi-kang;FANG Xi-bi;GAO Zhen;PAN Zi-yi;LIU Juan;JIANG Ping;YANG Run-jun;ZHAO Zhi-hui(College of Agriculture,Guangdong Ocean University,Zhanjiang,Guangdong 524088,China;College of Animal Science,Jilin University,Changchun 130062,China)
出处
《中国兽医学报》
CAS
CSCD
北大核心
2020年第3期568-574,共7页
Chinese Journal of Veterinary Science
基金
国家自然科学基金资助项目(31802034,31772562)
转基因生物新品种培育国家重点专项(2016ZX08009003-006)。
