摘要
为了研究羊口疮病毒(ORFV)F1L基因,以该病毒基因组DNA为模板,应用聚合酶链式反应(PCR)扩增得到1011bp的F1L基因,将其插入pMD18-TEasy克隆载体,构建了F1L基因克隆重组质粒;再将该基因片段插入pGEX-6P-1表达载体,通过PCR、限制性内切酶双酶切和测序鉴定,得到F1L基因插入方向和读码框结构均正确的阳性质粒,表明,成功构建了羊口疮病毒F1L基因的重组表达载体。通过对F1L基因序列进行生物信息学分析,预测发现F1L蛋白亲水、无信号肽,B细胞表位主要位于第4～8位、第16～22位、第32～53位、第59～73位、第82～99位和第123～133位氨基酸。这将为建立ORFV诊断方法、制备单克隆抗体及合成肽疫苗奠定基础。
In order to study F1L gene of orf virus（ORFV） ,the 1011bp-F1L gene of ORFV was amplified by PCR,and the PCR product was cloned into pMD18-T Easy vector.Then the purified F1L gene was subcloned into the pGEX-6P-1 vector and the recombinant plasmid was identified by PCR and restriction enzyme analysis.The authenticity and orientations of the gene were confirmed by sequencing.Results showed that the recombinant expression vector containing the ORFV F1L gene was successfully constructed.Bioinformatics analysis of the F1L gene showed that the F1L protein had hydrophilicity without signal peptide,and its B cell epitopes were located in 4 to 8,16 to 22,32 to 53,59 to 73,82 to 99 and 123 to 133 amino acids.The study laid the foundation for the development of methods for detection of ORFV,the preparation of monoclonal antibody against ORFV and the development of peptide vaccines against orf.
出处
《中国兽医科学》
CAS
CSCD
北大核心
2010年第11期1101-1105,共5页
Chinese Veterinary Science
基金
国家现代肉羊产业技术体系项目(nycytx-39)
