作 者：
左玉柱;乔木;王学理;向华;黄海楠;常爽;丁壮
单 位：
解放军军需大学军事兽医系;沈阳市东陵区动物防疫站
关键词：
鹅源副粘病毒;F蛋白基因;RT-PCR;re-Bacmid
摘 要：
鹅源副粘病毒分离株 NA- 1经 1 1日龄鸡胚增殖后收集尿囊液 ,浓缩 ,提取病毒基因组总 RNA,采用 RT- PCR方法 ,一次性扩增出与预期大小相符的特异性条带。将扩增产物提纯回收后克隆入 p MD1 8- T载体 ,经转化、筛选及酶切鉴定后 ,对阳性克隆进行了序列测定和序列分析。测序后拼接的 F基因长 1 6 72 bp,包含完整的开放阅读框 ( 1 6 6 2bp) ,编码 5 5 3个氨基酸 ,F蛋白裂解位点的氨基酸顺序为 1 1 2 R- R- Q- K- R- F1 1 7,与 NDV的强毒株特征相符 ,同时也与鹅副粘病毒分离株致病性试验结果相符。根据 NDV基因分型标准 ,鹅源副粘病毒 NA- 1株归属基因 型 NDV。将 F基因克隆入转座载体 p Fast Bac ,得到重组转座载体 PFF,将 PFF转化大肠杆菌 DH1 0 Bac,提取质粒 ,得到阳性重组穿梭载体 re- Bacm id,为下一步的转染表达奠定了基础
译 名：
Cloning of Fusion Glycoprotein Gene of Avian Paramyxovirus NA-1 Strain and Contructed of Re-bacmid
作 者：
ZUO Yu-zhu~1,QIAO Mu~2,WANG Xue-li~1,XIANG Hua~1,HUANG Hai-nan~1,CHANG Shuang~1, DING Zhuang~(1*) (1.Faculty of Military Veterinary,Quartermaster University of PLA,Changchun 130062, China;2.Dongling Epidemic Prevention Station of Shenyang,Shenyang 110015,China)
关键词：
geese′s paramyxovirus;F protein gene;RT-PCR;re-Bacmid
摘 要：
The geese′s paramyxovirus isolate NA-1 was propagated in 11-day-old chicken embryos.The allantonic fluid of the virus were concentrated and the genomic RNA was extracted.The F gene of the virus has been successfully amplified by RT-PCR,and further cloned into pMD18-T vector,and a positive recombitant plasmid was screened by restriction enzyme analysis.The sequence analysis showed that the nucleotide sequence of this F gene was 1 672 bp and encoding a protein of 553 amino acids.The amino acid sequence of cleavage site region was ~(112)R-R-Q-K-R-F~(117),matching to virulent NDV strains,and at the same time it was consistent with the pathogenicity test isolates.The geese′s paramyxovirus isolate NA-1 belongs to genotype Ⅶ.A recombinant transposon vector(pFF)was contructed by inserting the F gene of strain siping into the transposon vector pFastBacⅠ.Then,pFF was transformed into E.coli DH10Bac,and The recombinant bacmid was contructed.The recombinant bacmid can be used to transfect into sf-9 cells to generate recombinant baculovirus expressing F protein of NA-1.
