| 马氏珠母贝TLR6基因的克隆、序列分析与表达 |
| |
| 引用本文： | 吴羽媛,郭志颖,梁海鹰,林丽旋,郝瑞娟,焦钰.马氏珠母贝TLR6基因的克隆、序列分析与表达[J].水产学报,2017,41(11):1687-1698. |
| |
| 作者姓名： | 吴羽媛 郭志颖 梁海鹰 林丽旋 郝瑞娟 焦钰 |
| |
| 作者单位： | 1. 广东海洋大学水产学院,广东湛江 524025;广东省珍珠养殖与加工工程技术研究中心,广东湛江 524025;2. 广东海洋大学水产学院,广东湛江,524025 |
| |
| 基金项目： | 国家自然科学基金（31472306）；广东省科技计划项目（2012A031100010）；广东省海港建设与渔业产业发展专项（A201608B15）；国家级大学生创新创业训练项目（201410566005）；广东海洋大学大学生创新创业训练项目（CXXL2014012） |
| |
| 摘 要： | 为了探究TLR6(Toll like receptor 6)在马氏珠母贝免疫反应中的作用,实验采用c DNA末端快速扩增(RACE)技术获得了马氏珠母贝TLR6基因(Pm-TLR6)c DNA全长序列,并且运用实时荧光定量PCR(q RT-PCR)技术检测了Pm-TLR6在马氏珠母贝各组织中的表达情况、哈维氏弧菌刺激后和植核移植后血淋巴中的表达模式。结果显示:Pm-TLR6c DNA全长2295 bp,其中5′非编码区(UTR)长94 bp,3′UTR长89 bp,开放阅读框(ORF)长2112 bp,编码703个氨基酸;多序列比对结果表明物种间TLR6具有较高的保守性;PmTLR6具有跨膜域、富含亮氨酸的重复序列(LRRs)和TIR域,符合TLRs家族特征。q RTPCR数据分析表明,Pm-TLR6在马氏珠母贝肝胰脏、血细胞、鳃、性腺、闭壳肌、外套膜中均有表达,其中肝胰脏中表达量最高;哈维氏弧菌刺激后,Pm-TLR6在2 h表达上调,约为对照组(0 h)的9倍,随后的6 h恢复至正常水平,直至16 h表达水平开始回升并于24 h达到最大表达量,是对照组的29.4倍,具有显著性差异;植核移植实验结果显示PmTLR6在植核后5和10 d表达水平没有显著性变化,15和20 d表达量出现上升趋势,但差异不显著,最后于30 d达到最大值,约为空白对照组(0 d)的5倍,具有显著性差异。研究表明,Pm-TLR6可能在马氏珠母贝免疫防御反应中担任着重要的角色。
|
| 关 键 词： | 马氏珠母贝 TLR6 基因克隆 表达分析 |
| 收稿时间： | 2016/11/24 0:00:00 |
| 修稿时间： | 2017/3/28 0:00:00 |
Cloning, sequence analysis and expression studies on PmTLR6 gene from Pinctada fucata martensii |
| |
| WU Yuyuan,GUO Zhiying,LIANG Haiying,LIN Lixuan,HAO Ruijuan and JIAO Yu.Cloning, sequence analysis and expression studies on PmTLR6 gene from Pinctada fucata martensii[J].Journal of Fisheries of China,2017,41(11):1687-1698. |
| |
| Authors: | WU Yuyuan GUO Zhiying LIANG Haiying LIN Lixuan HAO Ruijuan and JIAO Yu |
| |
| Institution: | Fisheries College, Guangdong Ocean University, Zhanjiang 524025, China;Pearl Breeding and Processing Engineering Technology Research Center of Guangdong Province, Zhanjiang 524025, China,Fisheries College, Guangdong Ocean University, Zhanjiang 524025, China,Fisheries College, Guangdong Ocean University, Zhanjiang 524025, China;Pearl Breeding and Processing Engineering Technology Research Center of Guangdong Province, Zhanjiang 524025, China,Fisheries College, Guangdong Ocean University, Zhanjiang 524025, China,Fisheries College, Guangdong Ocean University, Zhanjiang 524025, China;Pearl Breeding and Processing Engineering Technology Research Center of Guangdong Province, Zhanjiang 524025, China and Fisheries College, Guangdong Ocean University, Zhanjiang 524025, China;Pearl Breeding and Processing Engineering Technology Research Center of Guangdong Province, Zhanjiang 524025, China |
| |
| Abstract: | TLR6 (Toll like receptor 6) is a kind of pattern recognition receptors and plays an important role in resisting microorganism infection. To study the function of TLR6 in the immune response of P. fucata martensii, in this study, a full length of Pm-TLR6 was obtained using rapid amplification of cDNA ends (RACE) technology from P. fucata martensii. The expression patterns of Pm-TLR6 in all tissues and its sequential expression in the hemolymph after Vibrio harveyi stimulation and nucleus insertion operation were further detected by Quantitative Real-Time PCR technology. Results showed that the total length of Pm-TLR6 cDNA was 2295 bp, including a 5'' UTR of 94 bp, a 3'' UTR of 89 bp and an open reading frame (ORF) of 2112 bp which encodes 703 amino acids. Multiple sequence alignment indicated that TLR6 was highly conservative among species. The protein encoded by Pm-TLR6 has a transmembrane domain, several leucine rich repeats(LRRs) and a TIR domain, conforming to the characteristics of TLRs family. qRT-PCR data revealed that Pm-TLR6 was expressed in all tested tissues, including hepatopancreas, hemocytes, gill, gonads, adductor muscle and mantle, with the highest expression in hepatopancreas (P<0.05). After Vibrio harveyi injection, the expression level of Pm-TLR6 increased at 2 h (9 fold vs. control), then dropped to normal levels at 6 h and began to increase at 16 h, with the highest level of expression appearing at 24 h (29.4 fold vs. control, p<0.05). The result of nucleus insertion surgery showed that Pm-TLR6 expression level was not significantly changed at 5 d and 10 d, and began to increase at 15 d and 20 d (P>0.05). Its expression reached the maximum level at 30 d with significant difference (5 fold vs. control, p<0.05). These results indicated that Pm-TLR6 may play an important role in the immune defense reaction of P. fucata martensii. |
| |
| Keywords: | Pinctada fucata martensii TLR6 gene cloning expression analysis |
| 本文献已被 CNKI 万方数据 等数据库收录！ |
| 点击此处可从《水产学报》浏览原始摘要信息 |
| 点击此处可从《水产学报》下载免费的PDF全文 |
|
