一株H_5亚型禽流感病毒血凝素基因的克隆与序列分析

根据GenBank中发表的H5亚型禽流感病毒HA基因序列设计2对引物,用RT-PCR方法从禽流感病毒广东分离病毒株(A/Chicken/Guangdong/1997)中扩增HA基因cDNA片段,并将其克隆至pMD-18T载体进行核苷酸序列测定。结果表明:用2对引物所扩增的片段大小分别约为1300 bp和800 bp,经序列拼接获得的HA基因cDNA长度约为1601 bp,编码533个氨基酸,与国内己发表的11个代表株的核苷酸和氨基酸序列同源性为分别为96.9%~99.9%和86.5%~93.0%;HA基因编码的氨基酸序列的系统进化树也表明A/Chicken/Guang-dong/1997、A/Goose/Huadong/01/2000、A/Ck/Hk/37.4/2002、A/Chicken/Zhoukou/2/02、A/Duck/Guangxi/53/2002、A/Duck/Fujian/01/2002等毒株处于同一进化枝,亲缘关系较近;而与A/Silly/Chicken/Hongkong/SF189/01株处于不同进化枝,亲缘关系较远。

Cloning and Sequence Analysis of Hemagglutinin Gene of the H5 Subtype Avian Influenza Virus Isolate

According to the nucleotide sequence of Hemagglutinin gene(HA) of H5 subtype avain influenza virus(AIV) published in GenBank,2 pairs of primers were deviced,and the cDNA fragments of HA gene of a AIV strain(A/Chicken/Guangdong/1997)isolated from Guangdong province were amplified by RT-PCR.The fragments were respectivly cloned into pMD-18T vector,and the recombinant plasmids were sequenced.The results showed that the cDNA fragments of HA gene consisted of 1300bp and 800bp respectively,and the full length of HA gene of the AIV isolate was composed of 1601bp,coding for 533 amino acids.The neucleitide and amino acids sequence homology compared with 11 AIV strains isolated from other areas of China were found to be 96.9~99.9% and 86.5~93.0%respectively.The phylogenetic tree also indicated the 6 strains(A/chicken/Guangdong/1997、A/Goose/Huadong/01/2000、A/CK/HK/37.4/2002、A/Chicken/Zhoukou/2/02、A/Duck/Guangxi/53/2002、A/Duck/Fujian/01/2002) clustered in the same or neighboring branches.However,the strain(A/Silly/Chicken/Hongkong/SF189/01) was distant in the evolutional origin.