【作者】 丁壮； 金宁一； 王兴龙； 金扩世； 李太元； 吕杰； 米志强； 夏志平； 殷震；

【Author】 DING zhuang,JIN ningyi,WANG xinglong,JIN kuoshi,LI taiyuan,Lii jie,MI zhiqiang,YIN zheng (The Military Veterinary Institute,The Quartermaster University of PLA Changchun, 130062)

【机构】 解放军军需大学军事兽医研究所； 吉林省龙井市延边大学农学院动医系； 沈阳农业大学畜牧兽医学院97硕研；

【摘要】 新城疫病毒(NDV)四平株、昌黎株、青岛株、F48E8株分别接种9日龄SPF鸡胚增殖,蚀斑纯化(三代),生物学特性鉴定及结合基因序列分析,表明NDV四平株、昌黎株、青岛株均为强毒株。经差速、蔗糖密度梯度离心提纯病毒,分别提取RNA,利用一对特异性引物及RT—PCR方法,一次性扩增出NDV四平株、昌黎株、青岛株和F48E8株的全长HN基因,分别将该HN基因克隆入载体pKS(-)并进行测序。序列分析表明。这四个毒株HN基因核苷酸长度均为1713bp,编码571个氨基酸,其中四平株和F48E8株含有5个糖基化位点,青岛株和昌黎株含有6个糖基化位点,四平株含有12个半胱氨酸残基,而昌黎株、青岛株和F48E8株含有13个半胱氨酸残基。三个野生毒株与F48E8相比较,核苷酸序列同源性为85.7%～99.3%,推导的氨基酸序列同源性为89.5%～98.8%,其中NDV四平株与标准强毒F48E8同源关系最近,核苷酸同源性为99.3%,推导的氨基酸同源性为98.8%。更多还原

【Abstract】 The NDV strains Siping, Changli, Qingdao and F48E8 were grown in 9 days SPF chicken embryos respectively and the viruses were purified by plaque formation . The identification of biological character and analysis of gene sequence revealed that the NDV strains Siping, Changli and Qingdao were highly virulent strains. The viruses were purified by sucrose density gradient centrifugation and the genomic RNAs were extracted respectively. A pair of special oligonucleotide primers were generated and whole sequences of HN gene of NDV strains Siping, Changli,Qingdao and F48E8 were amplified by RT-PCR. The HN genes of NDV strains Siping, Changli,Qingdao and F48E8 were sequenced after being directly inserted in plasmid pKS ( -) . The results showed that the nucleotide sequence of HN gene of the four strains of NDV was 1713bp long encoding a protein of 571 amino acids. There are 5 glycosylation sites in NDV strains Siping and F48E8 . There are 6 glycosylation sites in NDV strains Qingdao and Changli. There are 12 cysteine residues in NDV strain Siping. There are 13 cysteine residues in NDV strains Changli , Qingdao and F48E8 . Compared three wild type highly virulent strains of NDV with F48E8,the homology of the nucleotide sequence is 85. 7%-99. 3% and that of the deduced amino acid sequence is 89. 5%-98. 8%. The homology analysis and phylogenetic tree showed the more relationship between the NDV strains Siping and F48E8.更多还原