摘要
通过PCR方法扩增猪圆环病毒2型(pcv-2)中的缺失核定位序列的ORF2基因,将其克隆到Bac-N-Blue杆状病毒表达系统的pMelBacB载体中,得到阳性重组质粒pMel-ORF2。将其与Bac-N-Blue DNA体外重组,在脂质体的介导下,转染处于对数生长期的Sf9细胞,获得重组杆状病毒并在Sf9细胞上表达重组蛋白。通过SDS-PAGE、Western-blot和间接免疫荧光法(IFA)鉴定表明,Cap蛋白在昆虫-杆状病毒系统获得了表达,为Cap蛋白作为pcv-2检测抗原进行猪圆环病毒特异性抗体的检测提供了物质基础。
The ORF2 gene of Porcine Circovirus type 2（PCV2） deleting N-terminal nuclear localization region was amplified by PCR and cloned into the donor vector pMelBacB of Bac-N-Blue Baculovirus Expression System,and the recombinant plasmid was verified.After homologous recombination of the plasmid with Bac-N-Blue linear DNA,the mixture was transfected into Sf9 insect cells by Cellfectin Reagent,and the recombinant Baculovirus was constructed,which expressed recombinant protein on Sf9 cell.The result of confirming by SDS-PAGE,Western-blot and indirect lmmunofluorescence,indicated the recombinant Cap protein was expressed by Baculovirus system.It provided a substance foundation to test the specific antibody of Porcine Circovirus with Cap protein as detection antigen.
出处
《东北农业大学学报》
CAS
CSCD
北大核心
2012年第3期36-41,共6页
Journal of Northeast Agricultural University
基金
国家十一五支撑项目(2009BADB4B01)
