摘要： 根据GenBank中收录的鸡传染性喉气管炎病毒(ILTV)GX基因核苷酸序列，设计并合成了1对引物，应用PCR技术扩增ILTV河南株(ILTV-CG)GX基因。将ILTV-CG接种10日龄鸡胚，提取ILTV基因组DNA进行PCR扩增。将回收的PCR产物与pGEM-T Easy载体进行连接，转化JM109感受态细胞，经蓝白斑筛选，对菌液PCR和酶切鉴定为阳性的菌株进行测序。测序结果表明，所克隆的GX基因全长为950 bp，含有一个开放阅读框，编码299个氨基酸的多肽。推导的氨基酸有个潜在的N-糖基化位点，有11个与二硫键形成有关的半胱氨酸。序列分析表明，克隆的ILTV-CG GX基因与GenBank中发表的ILTV-SA2株GX基因核苷酸同源性为97.2%，变异点为第12位插入1个碱基G，第136位又插入2个碱基C，从而引起该毒株GX基因推导的氨基酸序列在第33位插人1个亮氨酸，还存在多处氨基酸发生变化，氨基酸同源性为95.3%。GX基因的成功克隆为构建ILTV GX基因缺失的病毒活载体奠定基础。

关键词: 传染性喉气管炎病毒, GX基因, 聚合酶链反应, 序列分析

Abstract: One pair of primers was designed according to the nucleotide sequence of chicken infectious laryngotracheitis virus (ILTV) GX gene from Genbank. The ILTV-CG strain was inoculated into 10-day embryonated eggs, which were used to extract the genome DNA for PCR amplification. The PCR products were purified, and then cloned into pGEM-T Easy vector. By screening of blue-white spot, the plasmid PCR and enzyme digestion, positive strains were sequenced. The results indicated that the amplified GX gene fragment was 950 bp, including an open-reading frame and encoding 299 amino acid residues that contain four potential N-glycosalation sites and eleven cysteines. The sequence analysis results showed a 97.2% similarity between GX gene ILTV-CG and ILTV-SA2 and 95.3% between their putative amino acid sequences. This study paved the way for preparation of recombinant vaccine of ILTV.

Key words: Infectious laryngotracheitis virus, GX gene, Polymerase chain reaction, Sequence analysis

中图分类号: 

• S432.4+1