缢蛏天然抗性相关巨噬蛋白2(<i>Sc-Nramp</i>2)基因克隆及其在副溶血弧菌刺激下的表达分析

林德海; 刘晨珊; 董迎辉; 何琳; 林志华

doi:10.3969/j.issn.0253-4193.2017.08.007

缢蛏天然抗性相关巨噬蛋白2(Sc-Nramp2)基因克隆及其在副溶血弧菌刺激下的表达分析

doi: 10.3969/j.issn.0253-4193.2017.08.007

浙江万里学院浙江省水产种质资源高效利用技术研究重点实验室, 浙江宁波 315100

基金项目: 国家现代贝类产业技术体系项目（CARS-48）；浙江省重大科技专项（2016C12907-7）；国家水产种质资源平台项目（2015DKA30470）；宁波市科技富民项目（2015C1000）；浙江省重中之重学科学生创新项目（CX2015011）；浙江省教育厅一般项目（Y201329410）。

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出版历程
- 收稿日期: 2016-11-23
- 修回日期: 2017-04-13

Cloning of natural resistance associated macrophage protein2 (Sc-Nramp2)gene from sinonovacula constricta and the expression analysis under vibrio parahaemolyticus stimulation

Key Laboratory of Aquatic Germplasm Resource of Zhejiang, Zhejiang Wanli University, Ningbo 315100, China

摘要

摘要: 克隆得到缢蛏天然抗性相关巨噬蛋白2（Sc-Nramp2）基因的cDNA全长序列3 681 bp，该基因的开放阅读框有1 776 bp，编码591个氨基酸，预测分子量为65.86 kDa；其结构具有Slc11蛋白家族的典型特征，包括有10个典型的跨膜结构和2个糖基化位点。Sc-Nramp2基因3'-UTR有2个类似于脊椎动物Nramp2中铁反应控制蛋白结合位点；同源性分析表明，Sc-Nramp2和太平洋牡蛎Nramp2-like的同源性最高为71.6%。实时荧光定量PCR结果表明，Sc-Nramp2基因在闭壳肌、外套膜、肝胰腺、斧足、水管和鳃6个组织中均有表达，其中肝胰腺中的表达量最高，其次是鳃，与其他组织均有极显著性差异（P<0.01）；注射副溶血弧菌后，肝胰腺中Sc-Nramp2基因的表达量较对照组显著上调（P<0.01），且表达量呈现先上升后下降的趋势，在12 h时达到最大表达量，推测Sc-Nramp2基因参与了缢蛏非特异性免疫应答反应。
- 缢蛏 /
- Sc-Nramp2 /
- 基因克隆 /
- 副溶血弧菌 /
- 基因表达
Abstract: The cDNA sequence of natural resistance associated macrophage protein in Sinonovacula constricta (Sc-Nramp2) was cloned by SMART RACE techniques. The ORF of cDNA sequence, coding for a protein of amino acids, was 1 773 bp and the molecular weight was deduced for 65.86 kDa. Sc-Nramp2 protein has the typical structural features with Solute carrier family 11 member, with 10 typical transmembrane domains and 2 glycosylation site. In 3'-UTR, Sc-Nramp2 was the presence of two IRE which was similar to the vertebrate Nramp2. Homology analysis indicated that the identity of Nramp with Crassostrea gigas was 71.6%. Real-time quantitative RT-PCR analysis indicated that the mRNA was expressed in all the six tissues, including adductor muscle, mantle, hepatopancreas, foot, waterpipe and gill. Nramp was expressed with the highest expression level in the hepatopancreas, then the gill, which had extremely significant differences from other tissues(P<0.01). The expression of Nramp in the hepatopancreas injected with V. Parahaemolyticus increased significantly(P<0.01). It rose to maximum expression at the 12^th hour and returned to the initial level. These results indicated that Sc-Nramp2 is involved in innate immunity of this species.
- Sinonovacula constricta /
- Sc-Nramp2 /
- gene cloning /
- Vibrio Parahaemolyticus /
- gene expression

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