Ⅰ类新城疫病毒NDV08-004株病毒拯救体系的建立  

Establishment of Reverse Genetics System for Class Ⅰ NDV08-004 Strain

作　　者：陈云霞[1,2] 刘华雷[1] 管峰[1] 郑东霞[1] 赵云玲[1] 王志亮[1] 

机构地区：[1]中国动物卫生与流行病学中心,青岛266032 [2]广西大学,南宁530004

出　　处：《病毒学报》2012年第5期496-500,共5页Chinese Journal of Virology

基　　金：国家自然科学基金(编号:30901079)资助项目

摘　　要：根据Ⅰ类新城疫病毒NDV08-004基因组序列(GenBank登录号FJ794269),设计7对引物,采用RT-PCR将NDV08-004基因组分段扩增(片段A～G)并分别克隆入pGEM-Teasy载体,然后采用单一的酶切位点将7个片段依次亚克隆到转录载体pOLTV5中,构建了含NDV08-004全基因组cDNA的转录载体NDV08-004-pO。采用构建的三个辅助质粒(pCI-NP、pCI-P和pCI-L)与基因组NDV08-004-pO按照2:1:1:2的比例共转染BSR T7/5细胞,结果成功拯救出具有血凝性的NDV08-004,初代分离物血凝价为4.33log2±0.58。Ⅰ类NDV反向遗传操作体系的建立为开展Ⅰ类NDV的致病机制奠定了基础。Based on the genomic sequence of NDV08-004 strain （GenBank accession number FJ794269）, seven pairs of primers were designed to amplify the genomic fragments by RT-PCR and cloned into pGEM- Teasy vector. The fragments （named A to G） were sub-cloned into transcription vector pOLTV5 according to the universal RE site and the plasmid named NDV08-004-pO which contained the full length cDNA of NDV08-004 strain was constructed. Three helper plasmids （pCI-NP, pCI-P and pCI-L） together with NDV08-004-pO were co-transfected into BSR T7/5 ceils, and the transfection supernatant was inoculated into SPF embryonated eggs to rescue the virus. The virus was rescued successfully and identified by HA and RT-PCR and sequencing. The rescue system constructed in this study provided a good foundation for the further related research.

关 键 词：Ⅰ类新城疫病毒 病毒拯救 

分 类 号：S852.65[农业科学—基础兽医学]