文章摘要
陶筱帆,谢婷婷,李小霞,罗军涛,白雅静,韩兵社,张俊芳.低温驯化下斑马鱼LINE1的表达检测.渔业科学进展,2020,41(3):88-93
低温驯化下斑马鱼LINE1的表达检测
Expression of LINE1 in Zebrafish (Danio rerio) During Cold Acclimation
投稿时间:2019-02-26  修订日期:2019-03-14
DOI:
中文关键词: LINE1  低温  斑马鱼  ZF4细胞  启动子
英文关键词: LINE1  Low temperature  Danio rerio  ZF4 cells  Promoter
基金项目:
作者单位
陶筱帆 上海海洋大学 水产种质资源发掘与利用教育部重点实验室 上海 201306 
谢婷婷 上海海洋大学 水产种质资源发掘与利用教育部重点实验室 上海 201306 
李小霞 上海海洋大学 水产种质资源发掘与利用教育部重点实验室 上海 201306 
罗军涛 上海海洋大学 水产种质资源发掘与利用教育部重点实验室 上海 201306 
白雅静 上海海洋大学 水产种质资源发掘与利用教育部重点实验室 上海 201306上海海洋大学 水产科学国家级实验教学示范中心 上海 201306 
韩兵社 上海海洋大学 水产种质资源发掘与利用教育部重点实验室 上海 201306上海海洋大学 水产科学国家级实验教学示范中心 上海 201306上海海洋大学 中国科学技术部海洋生物科学国际联合研究中心 上海 201306 
张俊芳 上海海洋大学 水产种质资源发掘与利用教育部重点实验室 上海 201306上海海洋大学 水产科学国家级实验教学示范中心 上海 201306上海海洋大学 中国科学技术部海洋生物科学国际联合研究中心 上海 201306 
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中文摘要:
      长散布核元件-1(Long spread nuclear element-1, LINE1)是跳跃基因。前期比较基因组研究发现,南极鱼经历漫长的低温适应进化后,与南极圈外的同亚目鱼类相比较,在基因水平上LINE1的扩增效率高达8~300倍,但LINE1的扩增与鱼类抵御寒冷之间的关系尚未明了。本实验对斑马鱼(Danio rerio)胚胎成纤维细胞ZF4进行了不同时间梯度的低温处理(18℃、5 d和18℃、30 d),同时对斑马鱼成鱼也进行了不同时间的低温处理(10℃, 3 h、6 h、1 d、3 d、5 d)。采用RT-qPCR检测了LINE1的mRNA水平,并克隆了斑马鱼LINE1基因启动子区,利用Luciferase双荧光报告系统,在ZF4细胞中验证LINE1 5UTR在低温压力下的生物活性。结果显示,短时间低温处理下,ZF4细胞中LINE1 mRNA水平有所降低,而在长时间低温处理中,LINE1的mRNA水平显著升高。在成鱼中,短时间低温处理下,LINE1 mRNA水平降低;长期低温处理下,LINE1 mRNA水平显著升高。在ZF4细胞中发现,LINE1 5UTR具有生物活性。在低温处理(18℃,3 d)下,报告基因信号减弱,间接表明LINE1启动子活性减弱。研究结果表明,低温压力会影响LINE1在鱼类中的表达。本研究为进一步探究LINE1在鱼类适应低温环境中的作用机制奠定了基础。
英文摘要:
      The long-spread nuclear element-1 (LINE1) retrotransposon is a mobile element in genome. Previous comparative genomic studies found that Antarctic Notothenioid fish underwent a long low-temperature adaptation evolution, and compared with Notothenioid fish outside the Antarctic circle, LINE1 genes were duplicated by 8~300 fold. The link between this augmentation and the resistance of fish to cold is not known. In this study, zebrafish (Danio rerio) embryonic fibroblasts ZF4 were exposed to a low temperature (18℃) for 5 days and 30 days, and adult zebrafish were exposed to a low temperature (10℃) for 3 h, 6 h, 1 d, 3 d, and 5 d. The mRNA expression of LINE1 was examined using RT-qPCR. The promoter regions of zebrafish LINE1 gene were cloned and the biological activity of LINE1 5'UTR at low temperature were verified in ZF4 cells by using the dual-luciferase reporter system. The following results were obtained: (1) In ZF4 cells, LINE1 mRNA expression was decreased by short-term low temperature treatment, but was significantly increased by long-term low temperature treatment. (2) In adult fish, LINE1 mRNA expression was decreased by short-term low temperature treatment, but was significantly increased in long-term low temperature treatment. (3) The LINE1 5'UTR was found to be biologically active in ZF4 cells. (4) It was found that during low temperature treatment (18℃, 3 d), the reporter gene signal was weakened, which indirectly indicated that the LINE1 promoter activity was weakened. The results showed that low temperature stress affected LINE1 expression in fish, which presents a foundation for further study on the mechanism of action of LINE1 in fish adaptation to a low temperature environment.
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