Department of Aquatic Life Medicine, Pukyong National University, Nam-gu 599-1, Busan, South Korea.
Appl Microbiol Biotechnol. 2010 Jan;85(3):679-90. doi: 10.1007/s00253-009-2177-3. Epub 2009 Aug 15.
In the present study, we have developed short interfering RNA (siRNA) expression vector utilizing rock bream beta-actin promoter and examined the possible use for the inhibition of highly pathogenic fish virus, rock bream iridovirus (RBIV), replication in vitro. Initially, in order to express siRNA effectively, we added several modifications to wild-type rock bream beta-actin promoter. Next, we succeeded in knocking down the expression of enhanced green fluorescent protein reporter gene expression in fish cells using newly developed vector more effectively than the fugu U6 promoter-driven vector we described previously. Finally, we could observe that cells transfected with modified rock bream beta-actin promoter-driven siRNA expression vector targeting major capsid protein (MCP) gene of RBIV exhibited more resistance to RBIV challenge than other control cells. Our results indicate that this novel siRNA expression vector can be used as a new tool for therapeutics in virus infection in fish species.
在本研究中,我们利用牙鲆β-肌动蛋白启动子开发了短发夹 RNA(siRNA)表达载体,并研究了其在体外抑制高致病性鱼类病毒牙鲆虹彩病毒(RBIV)复制的可能性。最初,为了有效地表达 siRNA,我们对野生型牙鲆β-肌动蛋白启动子进行了几种修饰。接下来,我们成功地利用新开发的载体,比我们之前描述的河豚 U6 启动子驱动的载体更有效地敲低了鱼类细胞中增强型绿色荧光蛋白报告基因的表达。最后,我们观察到转染了靶向 RBIV 主要衣壳蛋白(MCP)基因的经修饰的牙鲆β-肌动蛋白启动子驱动的 siRNA 表达载体的细胞对 RBIV 挑战的抵抗力比其他对照细胞更强。我们的结果表明,这种新型 siRNA 表达载体可作为鱼类病毒感染治疗的新工具。