EDSV六邻体蛋白基因克隆与原核表达载体构建

Cloning of Hexon-Encoding Gene of Egg Drop Syndrome Virus and Construction of Prokaryotic Expression Plasmids of Hexon Gene

应用降落PCR技术扩增出减蛋综合征病毒六邻体蛋白基因并将其克隆至pMDT18载体上,经酶切、PCR鉴定及测序结果表明插入的片段为目的基因,全长2.733kb,共编码910个氨基酸。切下该目的基因定向克隆至pET28a质粒构建了六邻体蛋白基因原核表达载体pET28ahexon,经各种酶切、PCR鉴定及进一步测序后证明六邻体蛋白基因片段所插入的位置、大小、核苷酸序列和阅读框架正确无误,从而为下一步的表达及进一步阐明EDSV六邻体蛋白基因结构与功能的关系和减蛋综合征基因工程苗的研究奠定了良好基础。

Hexon protein gene of egg drop syndrome virus (EDSV) was amplified by touchdown PCR(TD-PCR) and cloned into pMD18-T vector,and then sequencing results showed that the inserted fragment was the hexon gene of EDSV.The target gene was 2.74 kb and encoding a protein of 910 amino acids. The hexon gene was inserted into pET-28a vector, resulting in the construction of pET28a-hexon prokaryotic expression plasmids. The sequence analysis showed that the nucleotide sequence of hexon gene was 2.733kb and encoding a protein of 910 amino acids.The pET28a-hexon recombinant plasmids were identified by PCR and digested with enzymes and sequenced to confirm its rightness. The results will lay a sound foundation for further expression of hexon gene and the gene engineering vaccine of egg drop syndrome virus.