CAJ | 학술논문

通过RT-PCR方法扩增上海地区兔出血症病毒(RHDV)衣壳蛋白基因(VP60).将其克隆入腺病毒表达载体系统中,构建成重组腺病毒质粒,用脂质体法转染HEK293细胞,得到重组病毒pAd-VP60,经RT-PCR、IFA、HA、SDS-PAGE和Western-blotting鉴定,结果表明重组VP60蛋白在HEK293细胞中获得表达.表达产物与铝胶佐剂混合后注射3月龄非免疫健康家兔,结果表明,免疫后21 d兔体内产生抗RHDV抗体,其HI效价可达2~6～2~7,能抵御HA≥2~(10)致死剂量RHDV强毒的攻击,保护率为100%.
통과RT-PCR방법확증상해지구토출혈증병독(RHDV)의각단백기인(VP60).장기극륭입선병독표체재체계통중,구건성중조선병독질립,용지질체법전염HEK293세포,득도중조병독pAd-VP60,경RT-PCR、IFA、HA、SDS-PAGE화Western-blotting감정,결과표명중조VP60단백재HEK293세포중획득표체.표체산물여려효좌제혼합후주사3월령비면역건강가토,결과표명,면역후21 d토체내산생항RHDV항체,기HI효개가체2~6～2~7,능저어HA≥2~(10)치사제량RHDV강독적공격,보호솔위100%.
The capsid protein VP60 gene of rabbit hemorrhagic disease virus was amplified by reverse transcript pol-ymerase chain reaction (RT-PCR) technique. The fragment was cloned into the system of adenovirus expression and trans-fected into HEK293 cells, and a recombinant adenovirus named pAd-VP60 was obtained. The expressed protein was identified by RT-PCR, IFA, HA, SDS-PAGE and Westeru-blotting. The recombinant protein of VP60 was expressed in HEK293 cells. The expressed protein was mixed with aluminum rubber adjuvant and applied to im-munize three-month-old rabbits without immunized RHD vaccine. The results indicated that the anti-RHDV antibod-ies were induced and developed after 21 d of immuniza-tion, with the HI titers of serum ranging from 2~6 to 2~7.When challenged with HA> 2~(10) units of infectious RHDV, the control rabbits died while the immunized animals all survived. The results suggested that the recombinant pro-teins expressed by recombinant adenovirus could be promising candidates for RHDV live vaccines.