杨宏伟;刘振兴;郝乐;马江耀;梁志凌;马艳平;曹俊明;

• 1:仲恺农业工程学院
• 2:广东省农业科学院动物卫生研究所广东省畜禽疫病防治研究重点实验室农业农村部兽用药物与诊断技术广东科学观测实验站
• 3:广东海洋大学

摘要(Abstract)：

在开展花鲈(Lateolabrax maculatus)抗病毒饲料评价而采用qPCR进行病毒增殖以及免疫基因表达分析中需要首先确定病毒感染后稳定的内参基因。本研究选取12个组织/细胞(中肠、肌肉、胃、脑、心、肝、鳃、头肾、后肾、胸鳍、脾和花鲈肾细胞),采用qPCR方法检测花鲈虹彩病毒(Lateolabrax maculatus iridovirus,LMIV)感染前后RNAPoLⅡ、HPRT、TUBA、ACTB、18S rRNA、B2M、RPL7及EF1A8个基因的Ct值,采用GeNorm、NormFinder、BestKeeper软件评估内参基因表达的稳定性。以筛选的内参基因为基础建立qPCR方法,在LMIV感染花鲈肾细胞系10、24、48、72、96、120 h后对病毒ORF92R、基因进行表达分析。GeNorm软件结果表明8个候选内参基因在感染LMIV后的花鲈不同组织/细胞中的表达稳定性为:RNAPoLⅡ=HPRT>RPL7>TUBA>ACTB>18SrRNA>B2M>EF1A。NormFinder软件结果显示18S rRNA在对照组花鲈不同组织/细胞中表达稳定性最高;LMIV感染后RPL7是表达最稳定的内参基因。BestKeeper软件结果表明RPL7、RNAPoLⅡ、HPRT为花鲈感染LMIV后最佳参考基因组合。病毒基因的表达分析结果显示,RPL7、RNAPoLⅡ作为筛选出的稳定内参基因在分析病毒基因表达中优于18S rRNA。RPL7、RNAPoLⅡ、HPRT可作为评价花鲈感染LMIV后免疫相关基因表达以及病毒增殖的最佳qPCR参考基因,应用于花鲈抗病毒饲料的分析。

关键词(KeyWords)： 花鲈;qPCR;花鲈虹彩病毒;内参基因;抗病毒饲料

Abstract:

Keywords:

基金项目(Foundation): 佛山市市院农业科技合作项目;; 广东省农业科学院动物卫生研究所创新基金项目[CX-CP-202004]

作者(Author): 杨宏伟;刘振兴;郝乐;马江耀;梁志凌;马艳平;曹俊明;

Email:

DOI: 10.13302/j.cnki.fi.2020.24.008

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