为探究sox9基因在大黄鱼性别决定与分化过程中的作用,利用RACE方法克隆得到大黄鱼sox9a和sox9b 2个基因,采用实时荧光定量PCR(qRT-PCR)分析其在雌雄个体不同组织和不同发育阶段的表达规律,并检测了其在17β-雌二醇处理后的遗传雄性和17...为探究sox9基因在大黄鱼性别决定与分化过程中的作用,利用RACE方法克隆得到大黄鱼sox9a和sox9b 2个基因,采用实时荧光定量PCR(qRT-PCR)分析其在雌雄个体不同组织和不同发育阶段的表达规律,并检测了其在17β-雌二醇处理后的遗传雄性和17α-甲基睾酮诱导处理后遗传雌性幼鱼中的表达变化。结果显示,大黄鱼sox9a c DNA全长2 442 bp(NCBI登录号:MH996431),包含476 bp 5′UTR、466 bp 3′UTR、1 500 bp ORF,编码499个氨基酸。大黄鱼sox9b cDNA全长为2 199 bp(NCBI登录号:MH996432),包含335 bp5′UTR、415 bp 3′UTR、1 449 bp ORF,编码482个氨基酸。qRT-PCR分析显示,sox9a基因主要在性腺、眼、脑、肝脏中表达,精巢中的表达量显著高于卵巢;sox9b在大黄鱼各组织中广泛表达,但以精巢中表达量最高,而卵巢中只有痕量表达。在鱼苗性腺发育初期,sox9a/b的表达量都维持在较低水平;随着鱼苗长大,sox9a/b的表达量在84 dph、123 dph 2次达到高峰,随后开始下降,10 mph后又逐渐上升。17β-雌二醇处理能够显著下调遗传雄性大黄鱼sox9a和sox9b基因在性腺中的表达,17α-甲基睾酮处理则显著上调遗传雌性大黄鱼sox9a和sox9b基因在性腺中的表达。研究表明,sox9a/b基因与大黄鱼性别发育和分化过程密切相关,对大黄鱼精巢的发育与维持起重要的调控作用,而且两个基因的功能可能具有一定程度的分化。展开更多
Elovl4 is a fatty acyl elongase which participates in the biosynthesis of long-chain polyunsaturated fatty acids(LCPUFA).Herein we isolated and functionally characterized the elovl4a gene in the large yellow croaker L...Elovl4 is a fatty acyl elongase which participates in the biosynthesis of long-chain polyunsaturated fatty acids(LCPUFA).Herein we isolated and functionally characterized the elovl4a gene in the large yellow croaker Larimichthys crocea,and investigated the regulatory effects of transcription factors Hnf4α,Lxrα,Pparαon the activity of elovl4a promoter.Tissue expression pattern revealed that elovl4a was widely expressed in several tissues and predominantly in brain.Heterologous expression in yeast showed that L.crocea Elovl4a could effectively elongate both C18 and C20 PUFA substrates to C22 fatty acid.In addition,the affinity of large yellow croaker Elovl4a to n-6 series fatty acids is weak and cannot prolong 18:2n-6 and 18:3n-6.Moreover,L.crocea elovl4a reporter activities were elevated by 1.31-,1.39-and 1.48-fold via over-expression of Lxrα,Pparαand Hnf4α,respectively.The above findings can enrich the knowledge of biosynthesis pathway of LC-PUFA in the large yellow croaker and contribute to elucidate the LC-PUFA anabolism mechanism in fish.展开更多
文摘为探究sox9基因在大黄鱼性别决定与分化过程中的作用,利用RACE方法克隆得到大黄鱼sox9a和sox9b 2个基因,采用实时荧光定量PCR(qRT-PCR)分析其在雌雄个体不同组织和不同发育阶段的表达规律,并检测了其在17β-雌二醇处理后的遗传雄性和17α-甲基睾酮诱导处理后遗传雌性幼鱼中的表达变化。结果显示,大黄鱼sox9a c DNA全长2 442 bp(NCBI登录号:MH996431),包含476 bp 5′UTR、466 bp 3′UTR、1 500 bp ORF,编码499个氨基酸。大黄鱼sox9b cDNA全长为2 199 bp(NCBI登录号:MH996432),包含335 bp5′UTR、415 bp 3′UTR、1 449 bp ORF,编码482个氨基酸。qRT-PCR分析显示,sox9a基因主要在性腺、眼、脑、肝脏中表达,精巢中的表达量显著高于卵巢;sox9b在大黄鱼各组织中广泛表达,但以精巢中表达量最高,而卵巢中只有痕量表达。在鱼苗性腺发育初期,sox9a/b的表达量都维持在较低水平;随着鱼苗长大,sox9a/b的表达量在84 dph、123 dph 2次达到高峰,随后开始下降,10 mph后又逐渐上升。17β-雌二醇处理能够显著下调遗传雄性大黄鱼sox9a和sox9b基因在性腺中的表达,17α-甲基睾酮处理则显著上调遗传雌性大黄鱼sox9a和sox9b基因在性腺中的表达。研究表明,sox9a/b基因与大黄鱼性别发育和分化过程密切相关,对大黄鱼精巢的发育与维持起重要的调控作用,而且两个基因的功能可能具有一定程度的分化。
基金the Natural Science Foun-dation of Fujian Province(No.2018J01450)the National Natural Science Foundation of China(No.U1705231)+2 种基金the National Marine Fisheries Industrial Technology System Post Scientist Project(No.CARS-47-G04)the Scientific Research Foundation of Jimei University(No.ZQ2019046)the Central Public-Interest Scientific Institution Basal Re-search&Key Laboratory of Sustainable Development of Marine Fisheries,Ministry of Agriculture,P.R.China,CAFS(No.2017HY-XKQ01).
文摘Elovl4 is a fatty acyl elongase which participates in the biosynthesis of long-chain polyunsaturated fatty acids(LCPUFA).Herein we isolated and functionally characterized the elovl4a gene in the large yellow croaker Larimichthys crocea,and investigated the regulatory effects of transcription factors Hnf4α,Lxrα,Pparαon the activity of elovl4a promoter.Tissue expression pattern revealed that elovl4a was widely expressed in several tissues and predominantly in brain.Heterologous expression in yeast showed that L.crocea Elovl4a could effectively elongate both C18 and C20 PUFA substrates to C22 fatty acid.In addition,the affinity of large yellow croaker Elovl4a to n-6 series fatty acids is weak and cannot prolong 18:2n-6 and 18:3n-6.Moreover,L.crocea elovl4a reporter activities were elevated by 1.31-,1.39-and 1.48-fold via over-expression of Lxrα,Pparαand Hnf4α,respectively.The above findings can enrich the knowledge of biosynthesis pathway of LC-PUFA in the large yellow croaker and contribute to elucidate the LC-PUFA anabolism mechanism in fish.
