H5N1亚型高致病性禽流感病毒A/duck/HuBei/49/05株反向基因操作系统的建立

Establishment of Highly Pathogenic H5N1 Avian Influenza Virus A/duck/HuBei/49/05 Base on Reverse Genetics System

水禽是禽流感病毒的天然储存库,为了研究高致病性禽流感病毒对水禽致病性的分子基础,研究构建了DKHB/49/05的8质粒反向遗传操作系统。用Trizol从含A/duck/HuBei/49/05株病毒的鸡胚尿囊液中提取总RNA,用RT-PCR扩增PB2、PB1、PA、HA、NP、NA、M、NS基因片段,将其分别克隆至pBD载体中。利用8质粒反向遗传操作系统,将重组的pBD质粒共转染293T细胞,成功拯救了该病毒,并命名为R-DKHB。

Waterfowl are the natural repository of avian influenza virus.To study the pathogenic molecular basis of highly pathogenic avian influenza from waterfow,8 recombinant PBD plasmids and rescued the recombinant virus were constructrd by reverse genetics techniques.The total RNA was extracted from the allantoic fluids of chick embryo with the infection of A/duck/HuBei/49/05 virus,PB2,PB1,PA,HA,NP,NA,M and NS were amplified by RT-PCR respectively,and they were cloned into PBD vector.Moreover,they also were transfected into 293 T cells.The result indicated that the recombinant virus was successfully rescued and named as R-DKHB.