摘要
根据牛副流感病毒3型(BPIV-3)内蒙09株(NM09)全基因组序列(GenBank登录号:JQ063064)设计特异性引物,利用RT-PCR方法扩增出牛副流感病毒3型分离株的核衣壳蛋白(N)基因,通过NheI和NotI限制性内切酶位点亚克隆至真核表达载体pcDNA3.1/Zeo(+),获得真核重组质粒pcDNA3.1-N。采用Superfect转染试剂将重组质粒转染至BSR细胞中,转染后的BSR细胞经间接免疫荧光试验和RT-PCR方法检测,重组质粒pcDNA3.1-N在BSR细胞中能正确表达N蛋白。本研究结果为BPIV-3新型疫苗的研制奠定基础。
The aim of this study is to construct an eukaryotic expression plasmid for nucleocapsid protein (N) gene of bovine parainfluenza virus type 3 ( BPIV-3 ). According to the complete sequence of BPIV-3 NM09 strain ( GenBank accession : JQ063064 ), a pair of primers was designed and synthesized. The N gene of BPIV3 was amplified by reverse transcription polymerase chain reaction (RT-PCR). The N gene was cloned into pcDNA3. 1 (+) vector using NheI and NotI, and thus the expression plasmid pcDNA3.1-N was successfully construc- ted. Then pcDNA3. 1-N was transfected into BSR cell using lipoplast mediate method and positive results were demonstrated by indirect im- munofluorescence assay (IFA) and RT-PCR. The results showed N gene of BPIV-3 was successfully expressed in BSR cell. The study pro- vided fundamental information for the further study on the prevention of BPIV3 infection and new vaccine development.
出处
《畜牧与兽医》
北大核心
2012年第11期1-4,共4页
Animal Husbandry & Veterinary Medicine
基金
国家"863"项目(2011AA10A213)
关键词
牛副流感病毒3型
真核表达
N基因
bovine parainfluenza virus type 3
eukaryotic expression
N gene