中国明对虾天门冬氨酸转氨酶基因的克隆及氨氮胁迫对其时空表达的影响  

Cloning and expression analysis of aspartate aminotransferase c DNA in Fenneropenaeus chinensis following ambient ammonia stresses

作　　者：李少飞[1,2] 何玉英[2] 李吉涛[2] 李健[2] 刘萍[2] 葛倩倩[2,3] 

机构地区：[1]大连海洋大学水产与生命学院,辽宁大连116023 [2]中国水产科学研究院黄海水产研究所,山东青岛266071 [3]中国海洋大学,山东青岛266071

出　　处：《中国水产科学》2014年第6期1125-1133,共9页Journal of Fishery Sciences of China

基　　金：国家自然科学基金项目(31172401);国家虾产业技术体系专项项目(CARS-47);山东省自主创新专项项目(2013CX-80202);公益性行业(农业)科研专项(201103034);国家863计划项目(2012AA10A409)

摘　　要：利用RACE技术克隆获得中国明对虾(Fenneropenaeus chinensis)天门冬氨酸转氨酶GOT基因(Fc GOT)。Fc GOT基因c DNA全长为1 910 bp,其中,开放阅读框1 284 bp,编码427个氨基酸。同源性分析表明,中国明对虾天门冬氨酸转氨酶GOT氨基酸序列与其他节肢动物高度保守,与克氏原螯虾(Procambarus clarkii)和桔粉蚧壳虫(Planococcus citri)的同源性分别为78%和73%。系统进化分析表明,Fc GOT基因氨基酸序列与克氏原螯虾GOT聚为一支。组织表达分析发现Fc GOT基因在肝胰腺、鳃、血细胞、肌肉、心脏、淋巴中均有表达,其中肝胰腺中表达量最高。氨氮胁迫后,荧光定量PCR分析结果表明,Fc GOT基因在肝胰腺和鳃组织中的表达与对照组相比具有显著差异(P<0.05),表明Fc GOT基因在氨氮代谢方面具有重要的作用,参与了中国明对虾机体的急性氨氮胁迫应答反应。Fenneropenaeus chinensis is an important mariculture species in China. In aquaculture environments ammo-nia is a common toxic substance. In recent years, higher frequencies of ammonia nitrogen toxicity in shrimps have been reported. Therefore, it is necessary to investigate ammonia metabolism by F. chinensis. As an important member of the AAT-like family, the enzyme aspartate aminotransferase （GOT） is involved in many aspects of ammonia metabolism including participating in inosine monophosphate transdeamination, and the urea and citric acid cycles. Therefore, de-tailed understanding of the regulation of GOT is of great significance. In this study, we successfully cloned the aspartate aminotransferase cDNA of F. chinensis （FcGOT）. The FcGOT cDNA, which was 1 910 bp in length, contained a 5′-untranslated region（UTR） of 83 bp, a 3′UTR of 543 bp, and an open reading frame （ORF） of 1 284 bp, encoded a 427 amino-acid polypeptide. FcGOT protein exhibited typical AAT-like family features, including a Lys catalytic residue and 10 pyridoxal-5′-phosphate binding sites suggesting that it belongs to the AAT-I superfamily. Homology analysis revealed that the amino acid sequence of FcGOT was highly conserved with its homologs of other arthropods. The si-milarities of FcGOT with the GOT of Procambarus clarkia and Planococcuscitri were 78% and 73%, respectively. Phylogenetic analysis showed that FcGOT was in the same branch as P. clarkia. Tissue expression analysis showed that FcGOT transcripts were detected in all tested tissues, including hepatopancreas, gill, muscle, hemocytes, heart and lymph. The highest expression level of FcGOT was in the hepatopancreas. To study the biological functions of FcGOT in F. chinensis, the hepatopancreas and gill were sampled 1, 6, 12, 24, 48 and 72 h after exposure to different concen-trations of ammonia nitrogen. The expression of the FcGOT gene was significantly different compared with the control group （P〈0.05）, and the expression conditions differed between hepa

关 键 词：中国明对虾 天门冬氨酸转氨酶 基因克隆 基因表达 

分 类 号：S945[农业科学—水产养殖]