新城疫病毒B_(95)株HN蛋白基因的克隆及序列分析

Cloning and sequence analysis of HN gene of NDV B_(95)strain

根据 NDV HN蛋白已知基因序列设计一对引物 ,以提取的澳大利亚布里斯班 NDV分离株 B95基因组 RNA为模板 ,利用 RT- PCR技术扩增得到一条约 1.9kb的条带。将扩增产物克隆到 PGEM- T- easy载体中 ,并对重组质粒进行酶切分析、PCR鉴定 ,结果证明我们得到了 HN基因的阳性重组子 ,随后采用 Sanger's双脱氧末端终止法对阳性重组子进行核苷酸序列测定 ,获得该基因全长序列 ,并进行同源性分析。 B95株与其他毒株核苷酸同源性为 95 .1%～ 87% ,氨基酸同源性为96 .1%～ 92 .1%。

According to published HN protein gene sequence of NDV, we designed and synthesiezed a pair of primers. Genetic RNA of NDV B (95)strain, which was isolated from Brisbane in Australia, was used as template from cDNA synthesis of HN gene. A 1.9 Kb fragment was amplified from B (95) strain and cloned into the PGEM-T-easy vector. The recombinant plasmid was proved to be true by enzyme and PCR. The nucleotide sequence of HN gene was obtained by sanger's sequencing technique. Compared HN gene sequence of B (95)strain with the corresponding sequence of other strains of NDV, the homology of the nucleotide sequence was 95.1%～87%, the homology of the protein sequence was 96.1%～92.1%.