中国动物传染病学报2018，Vol.26Issue(2)：15-21,7.
表达Strep-tag的重组H5N6亚型禽流感病毒反向遗传操作系统的建立
ESTABLISHMENT OF A REVERSE GENETICS SYSTEM FOR H5N6 AVIAN INFLUENZA VIRUS EXPRESSING STREP-TAG
摘要
Abstract
To establish a reverse genetics system of H5N6 Avian influenza virus with Strep-tag, the strain A/Goose/Hunan/109/2014 (H5N6,HN109 strain)was rescued carrying a Strep-tag in the NS1 protein. The whole genome including 8 segments of H5N6 AIV HN109 strain was amplified by RT-PCR and then cloned into pBD vector. The corresponding sequence at position 77-84 in NS1 was replaced by Strep-tag (WSHPQFEK) through site-directed mutagenesis. The resulting 8 recombinant plasmids were used for co-transfection into 293T cells. The supernatant of 293T cells was collected at 48 h post-transfection and inoculated into 9-day-old specific pathogen free (SPF) chicken embryonated eggs. Then Strep-bearing virus (rHN109-NS1-Strep) was rescued and identified by hemagglutination assay. Sequence analysis confirmed that the 8 gene fragments were consistent with the planned design. The rHN109-NS1-Strep showed indistinguishable replication level from the parental strain HN109 both in MDCK cells and mice. All of these results confirmed the success of the reverse genetics system of Avian influenza virus H5N6 with Strep-tag, which provided a technological platform for further study on the molecular pathogenicity, transmission and development of novel vaccines.关键词
禽流感病毒/H5N6亚型/Strep-tagKey words
Avian influenza virus/H5N6 subtype/Strep-tag分类
农业科技引用本文复制引用
周圆一,王正祥,汪亮,徐帅,曾巧英..表达Strep-tag的重组H5N6亚型禽流感病毒反向遗传操作系统的建立[J].中国动物传染病学报,2018,26(2):15-21,7.基金项目
中央级公益性科研院所基本科研业务费(1610312016020) （1610312016020）
