松江鲈(<em>Trachidermus fasciatus</em>)泛素结合酶E2-D2基因的分子克隆及组织表达分析

陈学昭; 张雷; 于珊珊; 毕彩红; 刘春影; 祝茜

doi:10.3969/j.issn.0253-4193.2015.10.013

松江鲈(Trachidermus fasciatus)泛素结合酶E2-D2基因的分子克隆及组织表达分析

doi: 10.3969/j.issn.0253-4193.2015.10.013

山东大学(威海) 海洋学院, 山东威海 264209

基金项目: 威海市科技攻关计划项目(0000413420608); 威海市科委项目(1070432121313)。

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- 收稿日期: 2015-03-25

Molecular cloning and expression analysis of ubiquitin-conjugating enzyme E2-D2 in roughskin sculpin,Trachidermus Fasciatus

School of Marine Science, Shandong University (Weihai), Weihai 264209, China

摘要

摘要: 泛素结合酶(UbcE2)是蛋白泛素化过程中所必需的酶,在泛素转移和底物的特异性识别方面发挥着重要的作用。本实验利用RACE技术克隆获得了全长为993 bp的松江鲈泛素结合酶E2-D2基因cDNA序列(命名为TfUbcE2-D2),其开放阅读框为444 bp,编码147个氨基酸。通过SMART预测得知,TfUbcE2-D2含有一个UBCc结构域。同源比对结果表明该基因与其他物种的同源性为92.31%。实时荧光定量PCR显示,TfUbcE2-D2广泛表达于松江鲈各组织,在肾脏中的相对表达量最高,其次为鳃。鳗弧菌(Vibrio anguillarum)刺激后,TfUbcE2-D2在血液、脾脏、肝脏和鳃中表达均上调,其中,脾脏和鳃中的表达量上调约40倍。由以上实验结果推测,TfUbcE2-D2可能参与松江鲈的先天免疫防御。
- 泛素结合酶 /
- 松江鲈 /
- 基因克隆 /
- 非特异性免疫
Abstract: Ubiquitin-conjugating enzyme is the second enzyme in the process of protein ubiquitination and plays an important role in ubiquitin transferring and substrate specific recognition. In this study,we cloned the full-length cDNA from the Roughskin sculpin,Trachidermus fasciatus by RACE,and named as TfUbcE2-D2. The cDNA of TfUbcE2-D2 is 993 bp,which contained an open reading frame (ORF) of 444 bp encoding a polypeptide of 147 amino acids. The deduced amino acid sequence had a domain called Ubcc,which was extremely conservative. The mRNA expression of TfUbcE2-D2 in healthy and Vibrio anguillarum challenge was detected by quantitative real-time PCR,results showed that TfUbcE2-D2 was expressed in all the detected tissues,mainly expressed in kidney and gill. The up-regulated of TfUbcE2-D2 post challenge with Vibrio anguillarum in blood,spleen,liver and gill indicated that it might be involved in the innate response of Roughskin sculpin.
- ubiquitin-conjugating enzyme /
- Trachidermus fasciatus /
- gene clone /
- innate immunity

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