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由于环境、气候的恶化以及疾病的侵害,全球两栖动物种群数量呈现出整体衰退的趋势,其中虹彩病毒属蛙病毒科的蛙病毒是不可忽略的原因之一。环介导等温扩增技术(loop-mediated-isothermal-amplification,LAMP)是一种新型的核酸扩增技术,具有特异性强、快速、简便等特点,本研究利用LAMP技术,开发了针对中国东北林蛙种蛙蛙病毒的快速检测方法,针对蛙病毒MCP基因设计出能特异识别靶序列上6个位点的4条特异性引物,在链置换聚合酶(Bst 酶)的作用下,62 ℃扩增30 min。通过对实验参数的优化,扩增后病毒的最低检出数可达到102个拷贝。该方法可用于东北林蛙养殖厂种蛙蛙病毒的临床快速检测,保证种蛙的存活质量和数量。

Virus infection consists of entry, synthesis of macromolecular components, virus assembly and release. Understanding of the mechanisms underlying each event is necessary for the intervention of virus infection in human healthcare and agriculture. Here we report the visualization of Singapore grouper iridovirus (SGIV) assembly in the medaka haploid embryonic stem (ES) cell line HX1. SGIV is a highly infectious DNA virus that causes a massive loss in marine aquaculture. Ectopic expression of VP88GFP, a fusion between green fluorescent protein and the envelope protein VP088, did not compromise the ES cell properties and susceptibility to SGIV infection. Although VP88GFP disperses evenly in the cytoplasm of non-infected cells, it undergoes aggregation and redistribution in SGIV-infected cells. Real-time visualization revealed multiple key stages of VP88GFP redistribution and the dynamics of viral assembly site (VAS). Specifically, VP88GFP entry into and condensation in the VAS occurred within a 6-h duration, a similar duration was observed also for the release of VP88GFP-containing SGIV out of the cell. Taken together, VP088 is an excellent marker for visualizing the SGIV infection process. Our results provide new insight into macromolecular component recruitment and SGIV assembly.