鲤耐低温候选基因CcSCD全长cDNA的克隆及功能预测  

Isolation and Characterization of Full Length cDNA Sequence of A Stearoyl-CoA Desaturase Gene Responding to Low Temperature from Common Carp(Cyprinus carpio)

作　　者：常玉梅[1] 徐丽华[1,2] 李世国[1] 唐然[1] 梁利群[1] 

机构地区：[1]中国水产科学研究院黑龙江水产研究所,黑龙江哈尔滨150070 [2]上海海洋大学水产与生命学院,上海201306

出　　处：《水产学杂志》2012年第3期1-6,共6页Chinese Journal of Fisheries

基　　金：国家重点基础研究发展计划(2010CB126305);农业部淡水水产生物技术与遗传育种重点实验室(2012NYBSJ-06);中央级公益性科研院所基本科研业务费专项资金项目(HSY201205)

摘　　要：硬酯酰辅酶A脱氢酶(Stearoyl-CoA Desaturase,SCD)是参与脂肪酸脱氢反应的限速酶和关键酶,此酶编码基因SCD对增加膜磷脂的不饱和脂肪酸成分,提高细胞膜在低温下的流动性发挥重要作用,可能是抗寒过程中的关键基因成员。本研究采用cDNA末端快速扩增技术(Rapid Amplifications of cDNAEnds,RACE)克隆了鲤(Cyprinus carpio)脑组织CcSCD基因的全长cDNA序列,采用实时荧光定量PCR技术检测该基因在低温(6℃)和常温(23℃)条件下的转录表达差异,对其蛋白编码序列的结构和功能进行了预测分析。结果显示,CcSCD基因cDNA全长2618bp,包含一个由324个氨基酸残基组成的长度为975bp的阅读框(ORF);蛋白分子进化树显示鲤的SCD和草鱼(Ctenopharyngodon idella)的SCD蛋白同源性最高,相似性达88%;实时荧光定量结果表明,低温刺激下(6℃),CcSCD基因表达水平是常温条件下(23℃)的13.9倍,差异非常显著(P<0.01)。本研究旨在为今后构建表达载体,通过遗传操作等研究手段鉴定其抗寒功能奠定基础。The gene of stearoyl-CoA Desaturase（SCD） as a rate limiting enzyme involved in desaturation of saturated fatty acids plays vital roles in maintaining membrane fluidity and is considered as a key candidate which helps to improve cold tolerance.A SCD cDNA from common carp（Cyprinus carpio）（CcSCD） brain at cooling temperature was characterized and the transcription expression profiles were examined in the common carp exposed to cold（6℃） and warm（23℃） temperatures using real time RT-PCR.A full-length CcSCD cDNA was shown to be comprised of 2618 bp,including an open reading-frame of 975 bp and deduced 324 amino acid residues.Phylogenetic analysis revealed that the SCD protein sequences of the common carp and grass carp（Ctenopharyngodon idella） were clustered together,sharing the highest level of similarity（88%）.The quantified results showed that under cooling temperature,CcSCD transcripts were significantly up-regulated（P〈0.01）,in comparison to transcripts of samples from normal temperature.The findings help to establish a transgene construction and to apply genetic manipulation to identify functions of CcSCD gene of fish during cold response in future.

关 键 词：鲤 硬酯酰辅酶A脱氢酶 低温 荧光定量PCR 

分 类 号：Q754[生物学—分子生物学]