Microarray and RT-PCR screening for white spot syndrome virus immediate-early genes in cycloheximide-treated shrimp
Microarray and RT-PCR screening for white spot syndrome virus immediate-early genes in cycloheximide-treated shrimp
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DOI：
10.1016/j.virol.2005.01.047
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发表时间：
2005-04-10
期刊：
影响因子：
3.7
通讯作者：
Lo, CF
中科院分区：
文献类型：
--
作者：
Liu, WJ;Chang, YS;Lo, CF
在这里，我们报告的第一次成功使用放线菌酮（CHX）作为一种抑制剂，以阻止从头病毒蛋白质的合成在WSSV（白色斑点综合征病毒）感染。60个候选IE（立即早期）基因被确定使用全球分析微阵列技术。RT-PCR结果显示，台湾分离株ORF 126、ORF 242和ORF 418对应的基因均为CHX不敏感基因，分别命名为ie 1、ie 2和ie 3。这些IE基因的序列也出现在已经测序的另外两个WSSV分离株中。在中国分离株中鉴定了三个相应的ORF，但在泰国分离株中预测只有一个对应于ie 1的ORF。在使用EGFP（增强型绿色荧光蛋白）作为报告基因的Sf 9昆虫细胞中的启动子活性测定中，ie 1显示出非常强的启动子活性，产生比昆虫Orgyia pseudotsugata multicapsid核型多角体病毒（OpMNPV）ie 2启动子更高的EGFP信号。(c)2005年爱思唯尔公司All rights reserved.
Here, we report for the first time the successful use of cycloheximide (CHX) as an inhibitor to block de novo viral protein synthesis during WSSV (white spot syndrome virus) infection. Sixty candidate IE (immediate-early) genes were identified using a global analysis microarray technique. RT-PCR showed that the genes corresponding to ORF126, ORF242 and ORF418 in the Taiwan isolate were consistently CHX-insensitive, and these genes were designated ie1, ie2 and ie3, respectively. The sequences for these IE genes also appear in the two other WSSV isolates that have been sequenced. Three corresponding ORF's were identified in the China WSSV isolate, but only an ORF corresponding to ie1 was predicted in the Thailand isolate. In a promoter activity assay in Sf9 insect cells using EGFP (enhanced green fluorescence protein) as a reporter, ie1 showed very strong promoter activity, producing higher EGFP signals than the insect Orgyia pseudotsugata multicapsid nuclear polyhedrosis virus (OpMNPV) ie2 promoter. (c) 2005 Elsevier Inc. All rights reserved.