中国水产科学2011，Vol.18Issue(1)：1-7,7.DOI:10.3724/SP.J.1118.2011.00001
草鱼呼肠孤病毒VP6蛋白与大肠杆菌LTB亚基植物融合表达载体的构建
Construction of plant-based fusion expression vector of the GCRV VP6 antigenic protein and Escherichia coli LTB protein
摘要
Abstract
Grass carp reovirus (GCRV) is the first aquatic animal virus isolated and well characterized in China,which causes severe hemorrhage in fingerlings and yearlings of grass carp (Ctenopharyngodon idellus) with above 90％ morbidity and leads to huge economic losses. GCRV has been classified to the genus of aquareoviruses and shares the typical physico-chemical properties and morphological characteristics with other aquareovirueses,which consists of a double-layered capsid in spherical structure, containing a genome composed of 11 segments of double-stranded RNA. GCRV has been considered to be the most pathogenic aquareovirus in fishes, therefore, it represents a model system for the studies of replication and pathogenesis of aquareoviruses both in vitro and in vivo. Currently, there are limited vaccines available for the prevention and control of the disease, moreover, none of them is commercialized and able to be delivered through oral administration. The application of plants as a production system for vaccines facilitates a novel and safe approach of vaccination. Besides their application in human beings, they can be used as oral veterinary vaccines in animals. It should be noted that several plantderived vaccines for different animals, utilizing various plant production systems, are under investigation.Although up to date not yet readily available, the development of these plant-based vaccines is in an advanced stage and shows great potential for use in future. Especifically in aquaculture, oral vaccination has multiadvantages, since it is a relatively effortless, cost effective and stress-free immunization method, which can be used at almost any age of fish.For the purpose to generate a plant-based vaccine against grass carp hemorrhage, here we presented the result of constructing a plant-based fusion expression vector, in which a gut adhesion molecule gene (LTB), a GCRV antigenic peptide coding gene (GCRV-VP6) and a green fluorescent protein gene (GFP) were included. In this study the specific primer pairs for amplifying the GCRV-VP6 gene and the Escherichia coli LTB gene were designed and synthesized based on the sequences of GCRV-VP6 gene and Escherichia coli LTB gene posted in GenBank. GCRV genome was extracted from the virus infected grass carp kidney cell line (CIK) and the GCRV VP6 coding region was amplified by RT-PCR. The LTB coding region was amplified from Escherichia coli H44815 genome. The PCR products were cloned into pCR2.1 vector respectively and then sequenced and subcloned into vector pCAMBIA 1302, which contained a green fluorescence protein gene marker in order to track the route of antigen uptake. This study has made a solid base for the innovation of edible transgenic plant vaccine against grass carp hemorrhage through oral immunization.关键词
草鱼出血病/呼肠孤病毒/VP6基因/LTB基因/植物表达载体分类
生物科学引用本文复制引用
周勇,曾令兵,范玉顶,罗晓松,徐进,肖艺..草鱼呼肠孤病毒VP6蛋白与大肠杆菌LTB亚基植物融合表达载体的构建[J].中国水产科学,2011,18(1):1-7,7.基金项目
农业部现代农业产业技术体系建设专项资金资助(nycytx-49-16) （nycytx-49-16）
农业部公益性行业科研专项(200803013). （200803013）
