引用本文:	王昊盛,宋鑫金,董文强,吴斌,陈永霞,卢玮筱,祁鹏志,郭宝英.厚壳贻贝(Mytilus coruscus)金属硫蛋白MT-10:cDNA克隆、结构分析及铜离子胁迫下的表达.海洋与湖沼,2017,48(4):864-869.

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*厚壳贻贝(Mytilus coruscus)金属硫蛋白MT-10:cDNA克隆、结构分析及铜离子胁迫下的表达*
王昊盛, 宋鑫金, 董文强, 吴斌, 陈永霞, 卢玮筱, 祁鹏志, 郭宝英
浙江海洋大学国家海洋设施养殖工程技术研究中心舟山 316022

摘要:

近年来我国沿海海域重金属污染日趋严重，软体动物尤其是双壳贝类响应重金属污染胁迫的研究发展成为目前的研究热点。金属硫蛋白是一类广泛存在于生物体内富含半胱氨酸（Cys）的小分子蛋白，具有结合金属离子的能力，能有效调节生物体细胞内的金属离子平衡，具有清除自由基，重金属解毒的功能，关于厚壳贻贝MT蛋白的研究尚未见报道。本文首次利用RT-PCR和cDNA末端快速扩增技术克隆了厚壳贻贝MT-10蛋白cDNA全长（Mc-MT-10），其包含101bp的5'UTR区，108bp的3'UTR区以及222bp的ORF区，编码73个氨基酸。经比对发现Mc-MT-10富含Cys，且具有9个软体动物中常见的CXC结构以及CKCXXXCXCX结构，系统进化树中同科或属中MT-10聚合成一个分支，显示出高度保守性。采用Q-PCR技术分析了MT-10 mRNA在Cu²⁺胁迫下厚壳贻贝外套膜及消化腺中表达变化，结果表明Mc-MT-10 mRNA在Cu²⁺胁迫下两种组织中的表达量急剧升高，说明Cu²⁺能够诱导Mc-MT-10 mRNA的表达，并且表现出诱导具有阈值性特点。研究结果可为深入研究厚壳贻贝MT蛋白的功能奠定基础，为厚壳贻贝抗重金属污染新品种选育提供数据支持。

关键词: 厚壳贻贝金属硫蛋白基因荧光定量PCR 铜胁迫

DOI：10.11693/hyhz20170300056

分类号:Q789;Q955

基金项目:浙江省公益项目，2017C32009号；浙江海洋大学科研启动资金项目，21105013615号。

附件

THE CDNA CLONING AND CHARACTERIZATION OF MYTILUS CORUSCUS METALLOTHIONEIN 10 AS WELL AS ITS MRNA EXPRESSION UNDER THE CU²⁺ STRESS

WANG Hao-Sheng, SONG Xin-Jin, DONG Wen-Qiang, WU Bin, CHEN Yong-Xia, LU Wei-Xiao, QI Peng-Zhi, GUO Bao-Ying

Zhejiang Ocean University, National Engineering Research Center of Marine Facilities Aquaculture, Zhoushan 316022, China

Abstract:

In this paper, a 413bp full-length cDNA sequence of metallothionein 10 from thick shell mussel was obtained with RT-PCR and rapid amplification of cDNA ends (RACE) technique (so called ‘Mc-MT-10’ thereafter). It consists of a 101bp 5'untranslated region (UTR), an 222bp open reading frame (ORF) and a 108bp 3'UTR. The translated protein is composed of 73 amino acids. The multiple alignments showed the abundant Cys residues, 9 conserved CXC domains and one CKCXXXCXCX domain, in addition, the Mc-MT-10 cluster into one branch with the counterparts from congeneric species in the phylogenetic tree, suggesting the high evolutionary conservation. The Q-PCR assays were employed to assess the relative expressive changes of Mc-MT-10 mRNA in mantle and digestive gland under the stress of copper ions. The results showed that the expression levels of Mc-MT-10 mRNA rose sharply under the stress of Cu²⁺, implying that the Mc-MT-10 could involve in the regulation of thick hell mussel against Cu²⁺stress.

Key words: Mytilus coruscus metallothionein Real-time fluorescence quantitative PCR Cu²⁺stress