【摘要】 采用 RT- PCR技术扩增了禽流感病毒 A/ Guangdong/ 3/ 96 (H5 N1) [GD3/ 96 ]神经氨酸酶 (NA)基因 ,并将其克隆到 p UC18质粒中进行测序。核苷酸序列测定结果为 :NA基因全长为 14 10 bp,共编码 4 6 9个氨基酸。从 p UCNA中切下 NA基因片段 ,将其亚克隆到质粒 p SY5 38的 Eco R 位点 ,将带有痘苗病毒启动子 P11的 L ac Z基因平端克隆到该质粒的 Sm a 位点 ,然后切下同时含有 NA及 L ac Z基因的片段 ,再亚克隆到禽痘病毒载体 p SY6 81的 Not 位点 ,经限制性内切酶分析、PCR鉴定等 ,证明含有禽流感病毒 NA基因的重组禽痘病毒转移载体已构建成功 ,从而为进一步筛选表达 NA蛋白的重组禽痘病毒及探讨该蛋白的免疫原性奠定了实验基础。更多还原

【Abstract】 The cDNA of NA gene of avian influenza A/Goose/Guangdong/3/96(H5N1)isolate was amplified by RT-PCR, then cloned into pUC18 and sequenced. The result of sequencing showed that 1410bp of NA cDNA covered the complete open frame, encoding 468 amino acid residues. To construct the transfer vector, NA gene derived from recombinant plasmid pUCNA was subcloned into EcoRⅠ site of pSY538, and LacZ gene promoted by p11 was cloned into SmaⅠ site of this plasmid. Both NA and LacZ gene were cloned into NotⅠ site of FPV vector pSY681. Identified by restriction enzyme analysis and PCR, the transfer plasmid containing NA gene was constructed.更多还原