斑节对虾天门冬氨酸转氨酶基因的克隆及氨氮胁迫条件下的表达分析

发布时间：2018-11-27 08:46

【摘要】：为了探索天门冬氨酸转氨酶(AST)在斑节对虾(Penaeus monodon)氨氮(NH3-N)解毒代谢中的作用,该研究利用RACE技术获得了斑节对虾AST基因(PmAST)的cDNA全长序列,进行了相关生物信息学分析,在此基础上采用荧光定量与氨氮胁迫实验的方法研究了PmAST基因在斑节对虾的不同组织以及不同浓度氨氮胁迫过程中差异表达情况。该序列全长1 957 bp,开放阅读框(ORF)为1 242 bp,3'非编码区(UTR)为584 bp,包括含有30个碱基的poly(A)尾,5'非编码区(UTR)为131 bp。ORF可编码413个氨基酸,预测分子量为45.852 kD,理论等电点为8.85。序列含有1个AAT-like超家族结构域、15个磷酸化位点和2个糖基化位点。PmAST的mRNA在斑节对虾各组织中都有表达,在肝胰腺中表达量最高,其次为鳃组织,在胃和脑组织中的表达量最低。96 h氨氮胁迫后荧光定量PCR分析结果表明,PmAST在肝胰腺和鳃组织中都具有不同程度的表达上调,显著高于对照组(P0.05)。研究结果表明斑节对虾的PmAST基因在氨氮胁迫条件下会出现表达上调,并且氨氮浓度越高其上调幅度也越大,所以PmAST参与了斑节对虾的急性氨氮胁迫应答。
[Abstract]:In order to explore the role of aspartate aminotransferase (AST) in the detoxification and metabolism of (Penaeus monodon) ammonia nitrogen (NH3-N) in Penaeus vannamei, the cDNA full-length sequence of AST gene (PmAST) was obtained by RACE technique. Based on the analysis of bioinformatics, the differential expression of PmAST gene in different tissues of Penaeus Monodon (Penaeus Monodon) and different concentrations of ammonia nitrogen stress was studied by fluorescence quantitative and ammonia-nitrogen stress methods. The total length of 1 957 bp, open reading frame (ORF) is 1 242 bp,3' non-coding region, (UTR) is 584 bp, including poly (A) tail containing 30 bases, (UTR) of 5 'noncoding region 131bp.ORF can encode 413 amino acids. The predicted molecular weight is 45.852 kD, and the theoretical isoelectric point is 8.85. The sequence contained a AAT-like superfamily domain, 15 phosphorylation sites and 2 glycosylation sites. MRNA of PmAST was expressed in all tissues of Penaeus vannamei, the highest in hepatopancreas, followed by Gill tissue. The results of fluorescence quantitative PCR analysis showed that the expression of PmAST in hepatopancreas and Gill tissues was up-regulated in varying degrees, which was significantly higher than that in the control group (P0.05). The results showed that the expression of PmAST gene in Penaeus Monodon was up-regulated under ammonia-nitrogen stress, and the higher the concentration of NH3-N was, the larger the up-regulation was. So PmAST was involved in the acute ammonia-nitrogen stress response of Penaeus Monodon.
【作者单位】：中国水产科学研究院南海水产研究所农业部南海渔业资源开发利用重点实验室;上海海洋大学水产与生命学院;
【基金】：现代农业(虾)产业技术体系建设专项资金(CARS-47) 广东省省级科技计划项目(2014B020202003) 广东省海洋与渔业科技推广专项(A201501A06) 海南省自然科学基金项目(313117) 深圳市生物产业发展专项资金现代农业生物产业推广扶持计划项目(NYSW20140331010053)
【分类号】：S917.4

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