摘要
根据GenBank发表的猪圆环病毒2型(PCV2)ORF2基因序列设计并合成引物,采用重叠延伸PCR(splice overlap extension PCR,SOE-PCR)定点诱变技术对ORF2基因进行定点突变。DNA测序结果表明,ORF2基因片段的372—378 bp处的碱基由TCTAGAT突变为ATTGGAT,从而破坏酶切位点XbaⅠ,成功获得ORF2的突变基因片段。进而将其连接到腺病毒真核表达载体pAD5-Blue中,成功构建真核表达载体,并获得PCV2的Cap蛋白表达产物,为PCV2新型疫苗研究奠定了重要基础。
Based on the ORF2 sequences of PCV2 available in GenBank,two pairs of primers were designed and site-directed mutagenesis method based on overlap extension PCR was used to introduce mutations in the ORF2 fragment.DNA sequencing showed that TCTAGAT of 372 to 378 bp sites had been changed into TATAACT from mutagenesis,the endonuclease sites XbaⅠwas destroyed,site-directed mutagenesis was successfully implemented.Eukaryotic expression vector（pAD5-Blue） was constructed and expressed biological activity protein of Cap.It laid the foundation for further study and played a great role for the PCV2 subunit vaccine research.
出处
《中国畜牧兽医》
CAS
北大核心
2011年第2期99-103,共5页
China Animal Husbandry & Veterinary Medicine
