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脊尾白虾E75基因克隆及E75、ECR和RXR在不同蜕皮分期的表达分析
许杨1, 王佳佳1, 葛倩倩1, 崔彦婷1, 马骊1, 李健2,3
1.农业农村部海洋渔业可持续发展重点实验室 中国水产科学研究院黄海水产研究所 青岛 266071;2.农业农村部海洋渔业可持续发展重点实验室 中国水产科学研究院黄海水产研究所 青岛 266071;3.青岛海洋科学与技术试点国家实验室海洋 渔业科学与食物产出过程功能实验室 青岛 266071
摘要:
本研究在克隆脊尾白虾(Exopalaemon carinicauda)E75基因(EcE75, GenBank No. KY471317)的基础上,探讨了其在蜕皮过程中的作用,同时也探讨了克隆脊尾白虾的蜕皮激素受体基因(EcECR)和维甲酸X受体基因(EcRXR)在不同蜕皮分期的表达特征。克隆的EcE75基因全长为3944 bp,包括2520 bp的开放阅读框(ORF)。该开放阅读框编码一个由839个氨基酸组成的蛋白质,分子量为92.307 kDa,理论等电点为7.48。EcE75蛋白包含1个C4锌指结构,1个配体结合结构域,并且有1个明显的跨膜螺旋。EcE75基因在进化上具有一定的保守性,与甲壳动物聚为一支,与三疣梭子蟹(Portunus trituberculatus)、黑背地蟹(Gecarcinus lateralis)、凡纳滨对虾(Litopenaeus vannamei)、中国明对虾(Fenneropenaeus chinensis)等亲缘关系最近。EcE75基因在脊尾白虾的各组织中均有表达,其中,眼柄中的表达量最高,卵巢次之。在不同蜕皮分期中,EcRXR与EcECR、EcE75的表达规律基本一致,生殖蜕皮前期和后期表达量都比生长蜕皮高。生长蜕皮和生殖蜕皮因为卵巢发育而存在明显不同,蜕皮激素对卵巢发育的刺激作用,通过EcECR、EcRXR和EcE75基因的表达上调可以体现出来。对EcECR、EcRXR和EcE75基因在不同蜕皮分期的表达研究,为了解虾蟹类蜕皮机制提供了参考信息。
关键词:  脊尾白虾  EcECR  EcRXR  EcE75  蜕皮期
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Cloning of E75 gene and expression analysis of E75, ECR and RXR during different molting stages of Exopalaemon carinicauda
XU Yang,WANG Jiajia,GE Qianqian,CUI Yanting,MA Li,LI Jian
1.Key Laboratory of Sustainable Development of Marine Fisheries, Ministry of Agriculture and Rural Affairs, Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Qingdao 266071;2.Laboratory for Marine Fisheries Science and Food Production Processes, Pilot National Laboratory for Marine Science and Technology (Qingdao), Qingdao 266071
Abstract:
Based on the cloning of Exopalaemon carinicauda E75 (EcE75, GenBank accession number: KY471317), we studied its role in molting and the expression pattern of EcECR and EcRXR during different stages of molting. The full-length cDNA of EcE75 is 3944 bp long, which contains an open reading frame (ORF) of 2520 bp. The protein encoded by EcE75 is composed of 839 amino acids. The estimated mass of the deduced amino acid sequence of EcE75 is 92.307 kDa, and the calculated theoretical isoelectric point (pI) is 7.48. The ZnF_C4 and the HOLI domains were detected in EcE75, and one distinct transmembrane helix was identified. The gene EcE75 is conserved during evolution. The phylogenetic tree constructed based on E75 sequences showed that EcE75 was clustered with that of the same clade of crustaceans. E. carinicauda has the highest homology with Fenneropenaeus chinensis, Portunus trituberculatus, Litopenaeus vannamei, and Gecarcinus lateralis. The tissue distribution analysis showed that EcE75 transcript could be detected in all tested tissues of E. carinicauda, and relatively abundant in eyestalk and ovary. The expression pattern of the genes EcRXR, EcECR, and EcE75 was consistent during different stages of molting. At the premoult and postmoult stages, the expression of genes of reproductive molting was significantly higher than that of growth molting. The results showed that there was significant difference between the growth and reproductive molting because of the ovary development. Furthermore, it revealed the effect of ecdysone stimulation on ovarian development through the upregulation of expression of the genes EcRXR, EcECR, and EcE75. The present study on the expression of the genes EcRXR, EcECR, and EcE75 at different stages of molting provides a theoretical basis and a scientific ground for further research on the mechanism of molting.
Key words:  Exopalaemon carinicauda  EcECR  EcRXR  EcE75  Molting stage