团头鲂(Megalobrama amblycephala)caspase3基因克隆及其在高温胁迫中的表达分析  

Cloning of caspase3 Gene in Megalobrama amblycephala and Analysis of Its Expression in High Temperature Stress

作　　者：孙盛明 朱健[2] 戈贤平[2] 石连玉[1] 程磊[1] 张武肖[3] 苏艳莉 

机构地区：[1]中国水产科学研究院黑龙江水产研究所,农业部淡水水产生物技术与遗传育种重点实验室,哈尔滨150070 [2]中国水产科学研究院淡水渔业研究中心,农业部种质淡水渔业与种质资源利用重点实验室,无锡214081 [3]南京农业大学无锡渔业学院,无锡214081

出　　处：《基因组学与应用生物学》2017年第10期4090-4099,共10页Genomics and Applied Biology

基　　金：国家“十二五”科技支撑计划(2012BAD25B07);江苏省自然科学基金面上项目(BK20151103);国家大宗淡水鱼类产业技术体系华东养殖岗位(CARS-46-14);央级公益性科研院所基本科研业务费专项资金(2015C06XK01);农业部淡水水产生物技术与遗传育种重点实验室开放课题(KF2016-02)共同资助

摘　　要：采用RACE技术克隆获得团头鲂caspase3基因cDNA序列全长,并对该序列进行分析。结果显示,团头鲂caspase3基因全长为2 077 bp,开放阅读框长837 bp,5'非编码区长155 bp,3'非编码区长1 085 bp,命名为Ma Casp3。推测该基因编码278个氨基酸,预测分子量为30.92 ku,理论等电点为6.15。同源性和系统进化分析发现,Ma Casp3基因与斑马鱼(Danio rerio)caspase3氨基酸相似性最高,与其他节肢动物caspase3家族基因聚为一类。荧光定量RT-PCR结果显示,Ma Casp3基因在肝脏中的相对表达量最高,在肌肉中表达量最低。与常温对照组相比,高温胁迫7 d后该基因在团头鲂肝脏中的表达量显著升高,表明Ma Casp3基因可能参与团头鲂高温胁迫的应答并引发细胞凋亡级联反应。此外,高温胁迫7 d与胁迫前比较团头鲂幼鱼肝脏组织呈现肝细胞肿大、混浊变性、细胞核溶解、肝细胞空泡化现象,因此实际生产中应密切关注养殖水体水温变化以减少温度应激导致的组织损伤。In this study, full-length c DNA in caspase 3 was obtained by RACE in bluntnose black bream（Megalobrama amblycephala） and the sequence was identified. The full-length c DNA of Ma Casp3 was 2 077 bp and contained an open reading frame of 837 bp, the length of 5＇ noncoding region was 155 bp and 3＇noncoding region was 1 085 bp, which encoded a 278 amino acid protein with a calculated molecular mass of 30.92 k D and isoelectric point of 6.15. Homology and phylogenetic analysis revealed that amino acid of the Ma Casp3 gene had the highest similarity with zebrafish（Danio rerio） caspase3, and was associated with Caspase3 family genes of other arthropod. Quantitative real time PCR assay indicated Ma Casp3 expression had the high level in liver and the lowest in muscle. Compared with control group, Ma Casp3 gene expression upon high temperature challenge increased obviously which showed caspase 3 might be involved in thermal stress defense of bluntnose black bream. Liver organizations microstructure observation showed the liver tissue displayed nuclear hypertrophy,hepato cellular vacuolation, cellular outline indistinguishable, cytolysis, when fish juveniles exposed to high temperature stress. Thus, we should pay attention to variation of temperature in aquaculture pond to reduce the stress in organizations.

关 键 词：团头鲂 高温 肝脏 CASPASE3 基因克隆 

分 类 号：S917.4[农业科学—水产科学]