特异性检测珍珠龙胆石斑鱼虹彩病毒病的核酸适配体的筛选与鉴定

引用本文: 余庆, 刘明珠, 肖贺贺, 易弋, 程昊, Putra Dedi-Fazriansyah, 黎思巧, 李鹏飞. 特异性检测珍珠龙胆石斑鱼虹彩病毒病的核酸适配体的筛选与鉴定[J]. 分析化学, 2020, 48(5): 650-661. doi: 10.19756/j.issn.0253-3820.191647 shu

Citation: YU Qing, LIU Ming-Zhu, XIAO He-He, YI Yi, CHENG Hao, Putra Dedi-Fazriansyah, LI Si-Qiao, LI Peng-Fei. Selection and Characterization of Aptamers for Specific Detection of Iridovirus Disease in Cultured Hybrid Grouper (Epinephelus Fuscoguttatus ♀×E. Lanceolatus ♂)[J]. Chinese Journal of Analytical Chemistry, 2020, 48(5): 650-661. doi: 10.19756/j.issn.0253-3820.191647 shu

特异性检测珍珠龙胆石斑鱼虹彩病毒病的核酸适配体的筛选与鉴定

1.
广西科学院, 广西北部湾海洋研究中心, 广西海洋天然产物与组合生物合成化学重点实验室, 南宁 530007;
2.
广西科技大学, 生物与化学工程学院, 广西糖资源绿色加工重点实验室, 柳州 545006;
3.
广西科学院, 广西近海海洋环境科学重点实验室, 南宁 530007;
4.
河南师范大学生命科学学院, 新乡 453007;
5.
印度尼西亚亚齐大学, 班达亚齐 999006
基金项目:
本文系国家自然科学基金项目（Nos.41706145，41966004）、广西重点研发计划项目（No.2018AB52003）、广西自然科学基金项目（Nos.2019GXNSFAA245054，AD19245022）和广西科学院基本科研业务费项目（Nos.2019YJJ1005，2018YJJ903，2018YJJ902）资助

摘要: 石斑鱼虹彩病毒是一种核质DNA病毒，不仅给海水养殖造成巨大的经济损失，而且严重威胁全球生物多样性。为了进一步阐明石斑鱼虹彩病毒侵染致病的分子机理，同时为石斑鱼虹彩病毒病的预防、诊断和治疗提供新的理论依据，本研究以珍珠龙胆石斑鱼虹彩病毒（SGIV-Gx）感染的宿主细胞为靶标，采用指数富集配体系统进化技术（SELEX）筛选获得4条核酸适配体（LYGV1、LYGV2、LYGV3、LYGV4），并对核酸适配体的性质进行了系统的分析研究。各条核酸适配体识别SGIV-Gx病毒感染的宿主细胞具有高特异性和高亲和性，筛选的核酸适配体无毒副作用，而且核酸适配体LYGV3具有一定的抗病毒效果。对核酸适配体的靶标性质进行了分析，结果表明，核酸适配体（LYGV1、LYGV2、LYGV3、LYGV4）的靶标可能是细胞膜蛋白或膜蛋白相关成分。其中，核酸适配体LYGV1的靶位点在SGIV-Gx病毒感染细胞2 h后出现在宿主细胞表面，核酸适配体LYGV2、LYGV3和LYGV4的靶位点的出现时间分别为病毒感染细胞后的4、8和6 h。

关键词:

English

Selection and Characterization of Aptamers for Specific Detection of Iridovirus Disease in Cultured Hybrid Grouper (Epinephelus Fuscoguttatus ♀×E. Lanceolatus ♂)

1.
Guangxi Key Laboratory of Marine Natural Products and Combinatorial Biosynthesis Chemistry, Guangxi Beibu Gulf Marine Research Center, Guangxi Academy of Sciences, Nanning 530007, China;
2.
Guangxi Key Laboratory of Green Processing of Sugar Resources, College of Biological and Chemical Engineering, Guangxi University of Science and Technology, Liuzhou 545006, China;
3.
Guangxi Key Laboratory of Marine Environmental Science, Guangxi Academy of Sciences, Nanning 530007, China;
4.
College of Life Science, Henan Normal University, Xinxiang 453007, China;
5.
Department of Aquaculture, Syiah Kuala University, Banda Aceh 999006, Indonesia
Received Date: 03 November 2019
Revised Date: 09 February 2020
Fund Project:
This work was supported by the National Natural Science Foundation of China (Nos. 41706145, 41966004), the Key Research and Development Programs of Guangxi Province, China (No. 2018AB52003), the Natural Science Foundation of Guangxi Province, China (Nos. 2019GXNSFAA245054, AD19245022), and the Basic Research Fund of Guangxi Academy of Sciences (Nos. 2019YJJ1005, 2018YJJ903, 2018YJJ902).

Abstract: Grouper iridovirus is a nucleocytoplasmic DNA virus, which not only causes great economic losses in mariculture but also seriously threatens the global biodiversity. To further elucidate the molecular mechanism of grouper iridovirus pathogenesis, and provide the new theoretical basis for the prevention, diagnosis and treatment of grouper iridovirus disease, in this study, we generated four single-stranded DNA aptamers against SGIV-Gx infected cells using systematic evolution of ligands by exponential enrichment (SELEX). Then, the properties of aptamers were characterized. Firstly, each aptamer exhibited high specificity and affinity to grouper iridovirus Guangxi strain (SGIV-Gx)-infected cells. Secondly, aptamers exerted no cytotoxic effects, and aptamer LYGV3 appeared to inhibit SGIV-Gx infection in vitro. The target molecules of LYGV1, LYGV2, LYGV3, LYGV4 could be directly or indirectly related to surface proteins anchored on the membranes of SGIV-Gx infected cells. The targets of aptamers (LYGV1, LYGV2, LYGV3, LYGV4) appeared on the cells surface at 2, 4, 8, and 6 hour post infection.

Key words:

Grouper iridovirus
/ Systematic evolution of ligands by exponential enrichment
/ Aptamer
/ Specificity
/ Antiviral activity
/ Target protein

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沈阳化工大学材料科学与工程学院沈阳 110142

特异性检测珍珠龙胆石斑鱼虹彩病毒病的核酸适配体的筛选与鉴定

English

Selection and Characterization of Aptamers for Specific Detection of Iridovirus Disease in Cultured Hybrid Grouper (Epinephelus Fuscoguttatus ♀×E. Lanceolatus ♂)

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