P基因对小反刍兽疫病毒基因组转录和复制的分子调控机制研究

Peste des petits ruminants (PPR) is an acute and highly contagious viral disease of small ruminants causing high morbidity rates and sometimes high mortality rates. Since the first outbreak of PPR in Tibet in 2007, PPR has seriously affected small ruminant production in Tibet. The phosphoprotein of paramyxoviruses is known to play multiple roles in both transcription and replication, being an indispensable subunit of the viral polymerase complex. The objective of this project is to study the role of phosphoprotein of PPRV in viral RNA synthesis in virus infection using reverse genetics. Minigenome system on virulent PPRV strain Tibet 07 will be constructed. Using this minigenome system, the P gene will be exchanged between the attenuated PPRV vaccine strain Nigeria 75/1 and the virulent PPRV strain Tibet 07. Mutations of key amino acid of P of PPRV Tibet 07 will be constructed. The activities of the mutant P proteins in the minigenome system will be determined. Furthermore, we aim to rescue live PPR virus from a full-length cDNA copy of the genome of the Tibet 07 strain. The recombinant will be constructed in which P gene is replaced that from Nigeria 75/1 or mutated. The recombinant virus will be rescued and the viral growth will be examined in cell culture monolayers. This study will expand our knowledge on the role of phosphoprotein in genome transcription and replication of PPR virus. This study will offer a basis on investigating the pathogenicity of PPRV.

小反刍兽疫是由小反刍兽疫病毒引起的一种感染山羊和绵羊的严重的烈性、接触性传染病。小反刍兽疫曾经给我国西藏养羊业造成巨大经济损失，目前仍然威胁着我国西南边境省份的家畜生产安全。研究小反刍兽疫病毒感染致病机制，对于指导疫病的预防和控制具有重要的意义。小反刍兽疫病毒P基因对于病毒基因组转录和复制至关重要。本课题拟利用反向遗传操作技术研究小反刍兽疫病毒强毒株Tibet 07的P基因在病毒RNA合成中的作用。首先，构建小反刍兽疫病毒强毒株Tibet 07的微型基因组，进行P基因替换和定点突变，比较微型基因组活性变化。其次，构建Tibet 07株的感染性全长cDNA克隆并拯救病毒，进行P基因替换和定点突变，研究重组病毒基因组转录和复制水平变化。通过上述研究探讨P基因对病毒基因组转录和复制的调控作用，探求关键的作用位点，为进一步开展小反刍兽疫的感染致病机制研究奠定必要的基础。

小反刍兽疫是由小反刍兽疫病毒引起的一种感染山羊和绵羊的严重的烈性、接触性传染病。小反刍兽疫曾经给我国西藏养羊业造成巨大经济损失，目前仍然威胁着我国西南边境省份的家畜生产安全。研究小反刍兽疫病毒感染致病机制，对于指导疫病的预防和控制具有重要的意义。小反刍兽疫病毒P基因对于病毒基因组转录和复制至关重要。本课题拟利用反向遗传操作技术研究小反刍兽疫病毒强毒株Tibet 07的P基因在病毒RNA合成中的作用。首先，构建了小反刍兽疫病毒强毒株Tibet 07的微型基因组，能够表达外源性绿色荧光蛋白。其次，构建了Tibet 07株的全长cDNA克隆。最后，在昆虫细胞中表达了Tibet 07株的P蛋白，鉴定具有抗原性。上述研究为探讨P基因对病毒基因组转录和复制的调控作用，探求关键的作用位点，并进一步开展小反刍兽疫的感染致病机制研究奠定必要的基础。