对虾白斑综合征病毒wsv477基因的克隆表达与时相分析

Clone and expression and temporal analysis of wsv477 gene from white spot syndrome virus

将对虾白斑综合征病毒(white spot syndrome virus,WSSV)的功能基因wsv477克隆到原核表达载体PGEX-4T-2中,并转化大肠杆菌BL21,37℃条件下诱导了WSV477蛋白的表达,并以纯化的融合蛋白作为抗原,免疫小鼠获得了多克隆抗体.该基因开放阅读框共624个核苷酸,编码208个氨基酸,GST融合蛋白相对分子质量为49000,抗体滴度为1:10000.RT-PCR和Western blot分析表明,该基因在病毒感染4h后开始转录,6h后开始表达,是WSSV感染的早期基因.

The wsv477 gene from WSSV was cloned in pGEX-4T vector and expressed in Escherichia coli BL21.Then the wsv477 protein was induced and expressed at 37 ℃,and the purified fusion protein was used as antigen to immunize mice,subsequently the specific polyclony antibody was raised using the purified fusion protein（GST-WSV477）.The ORF of wsv477 gene had 624 base pairs and encoded 208 amino acid peptides,the GST fusion protein was 49 000,and the titer of the antisera was 1：10 000.RT-PCR and Western blot revealed that the wsv477 gene was first transcribed at 4 h postinfection,and was expressed at 6 h postinfection in vivo,suggesting that it was an early gene.