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当前位置: 首页 > 文章 > 建鲤极长链脂酰辅酶A脱氢酶基因cDNA克隆及相关定量表达分析 南方农业学报 2015,46 (4) 680-686
Position: Home > Articles > Cloning,sequence analysis and relevant quantitative expression analysis of VLCAD cDNA from Cyprinus carpio var. Jian Journal of Southern Agriculture 2015,46 (4) 680-686

建鲤极长链脂酰辅酶A脱氢酶基因cDNA克隆及相关定量表达分析

作  者：
刘骏恂;俞菊华;李红霞;唐永凯;李建林;于凡
单  位：
中国水产科学研究院淡水渔业研究中心/农业部淡水鱼类遗传育种重点实验室;南京农业大学无锡渔业学院
关键词：
建鲤;VLCAD基因;组织表达量;脂肪源;饥饿胁迫
摘  要：
【目的】克隆建鲤(Cyprinus carpio var.Jian)极长链脂酰辅酶A脱氢酶(VLCAD)基因c DNA序列,并分析其表达情况,为揭示鲤鱼脂肪酸代谢机理提供理论依据。【方法】以建鲤为研究对象,利用RT-PCR、RACE克隆VLCAD基因的c DNA序列,并通过实时荧光定量PCR对VLCAD基因在不同组织、脂肪源及饥饿胁迫等条件下的表达情况进行分析。【结果】扩增获得的建鲤VLCAD基因c DNA序列全长2527 bp,包括296 bp的5'非翻译区、1974 bp开放阅读框(OFR)及257 bp的3'非翻译区,共编码659个氨基酸;建鲤VLCAD氨基酸序列包含FAD结合位点、底物结合位点、催化位点,具有ACADs家族的特征性结构;与鲤科类斑马鱼(Danio rerio)的同源性最高,达94%。建鲤VLCAD基因在性腺、肌肉、肝脏、前肠、肾脏、心脏和脑中均有表达,且以心脏的表达量最高,前肠的表达量最少。鱼油和亚麻油对建鲤VLCAD基因在肝脏和肌肉中的表达无显著影响(P>0.05),但饥饿状态下肌肉中的VLCAD基因表达量显著高于正常状态(P<0.05)。【结论】建鲤VLCAD基因在富含线粒体且脂肪酸代谢旺盛的组织器官中高效表达,对脂肪酸β氧化起调控作用,尤其在饥饿状态下对脂肪酸氧化调控作用明显增强,能促进脂肪酸分解为机体供能。
译  名：
Cloning,sequence analysis and relevant quantitative expression analysis of VLCAD cDNA from Cyprinus carpio var. Jian
作  者：
LIU Jun-xun;YU Ju-hua;LI Hong-xia;TANG Yong-kai;LI Jian-lin;YU Fan;Wuxi Fishery College,Nanjing Agricultural University;Freshwater Fisheries Research Center,Chinese Academy of Fishery Sciences/Key Laboratory of Freshwater Fisheries and Germplasm Resources Utilization,Ministry of Agriculture;
关键词：
Cyprinus carpio var.Jian;;VLCAD gene;;tissue expression;;lipid source;;starvation stress
摘  要：
【 Objective 】 The sequence of very-long-chain acyl-Co A dehydrogenase( VLCAD) of Cyprinus carpio var. Jian was cloned and its expression was studied to provide references for revealing fatty acid metabolism mecha-nism of Cyprinus carpio var. Jian. 【Method 】A full-length VLCAD c DNA was obtained and sequenced from Cypri nus carpio var. Jian by utilizing RT-PCR and RACE methods. Real-time fluorescence quantitative PCR was used to get the expression of jl-VLCAD in different tissues, lipid sources and starvation stress. 【Result】The jl-VLCAD was 2527 bp in length, containing 296 bp 5′-untranslated region, 257 bp 3′-untranslated region and 1974 bp open reading frame encoding 659 amino acids. Flavin Adenine Dinucletide binding site, substrate binding pocket and catalytic base were consistent with its family characteristic structure. The results of semi-quantitative RT-PCR indicated that the jl-VLCAD m RNA was transcribed ubiquitously in all seven adult tissues( gonad, muscle, liver, foregut, kidney, heart,brain), with expression being highest in the heart, and lowest in foregut. Comparing with the expressions of jl-VLCAD in two groups of lipid sources, there was no significant difference(P>0.05)between fish oil group and linseed oil group.After two weeks starvation stress, the expression of jl-VLCAD significantly increased in muscle(P<0.05). 【Conclu sion 】The expression of jl- VLCAD is quite high in tissues rich in mitochondrion and fatty acid metabolism. It plays an important role in regulation of the β-oxidation pathway of fatty acids, and the effect of jl-VLCAD on the oxidation of fatty acid regulation is obviously strengthened for energy supply after starvation stress.

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