CAJ | 학술논문

采用RT-PCR方法从传染性造血器官坏死病病毒( IHNV)中扩增RNA获得1176 bp的核衣壳蛋白( N)基因,将其插入于pFB-LIC-Bse杆状病毒载体中,构建了重组质粒pFB-LIC-Bse-N,转化至感受态细胞DH10Bac中,获得重组杆粒rBacmid-N。再将其转染至Sf9昆虫细胞,获得重组杆状病毒。 IFA和Western Blot分析表明,重组N蛋白可以被辣根过氧化物酶(HRP)标记的组氨酸单抗(anti-His)识别,表明,IHNV N基因在昆虫细胞Sf9中获得了正确表达。为深入研究N蛋白的功能和免疫学特性奠定了基础。
채용RT-PCR방법종전염성조혈기관배사병병독( IHNV)중확증RNA획득1176 bp적핵의각단백( N)기인,장기삽입우pFB-LIC-Bse간상병독재체중,구건료중조질립pFB-LIC-Bse-N,전화지감수태세포DH10Bac중,획득중조간립rBacmid-N。재장기전염지Sf9곤충세포,획득중조간상병독。 IFA화Western Blot분석표명,중조N단백가이피랄근과양화물매(HRP)표기적조안산단항(anti-His)식별,표명,IHNV N기인재곤충세포Sf9중획득료정학표체。위심입연구N단백적공능화면역학특성전정료기출。
To study the major structural nucleoprotein ( N ) of infectious hematopoietic necrosis virus ( IHNV ) by baculovirus systems,in this study nucleoprotein ( N) gene(1 176 bp) was amplified by RT-PCR from infectious hematopoietic necrosis virus ( IHNV) and inserted into the baculovirus vector pFB-LIC-Bse to construct a recombi-nant plasmid pFB-LIC-Bse-N. The constructed recombinant transposition plasmid pFB-LIC-Bse-N was transformed to competent E. coli DH10Bac to get recombinant bacmids rBacmid-N. The obtained recombinant bacmid rBacmid-N was transfected to insect Sf9 cells,and the recombinant baculovirus that contained N gene was obtained. IFA and Western Blot analysis showed that the recombinant N protein can be recognized by horseradish peroxidase ( HRP) labeled histidine monoclonal antibody ( anti-His) which indicated IHNV N gene has been successfully expressed in Sf9 cells infected with recombinant baculovirus. The study laid a foundation of further study on N protein structure, function and immunological characteristics.