【摘要】 为构建表达鸡新城疫病毒(NDV)F基因的重组鸭肠炎病毒(DEV),本研究以实验室前期构建的表达绿色荧光蛋白的重组DEV(r DEV-US10-EGFP)为基础,通过PCR扩增NDV F基因,构建了携带F基因表达盒的转移质粒p US10-TPA-F。将转移质粒与r DEV-US10-EGFP基因组DNA共转染鸡胚成纤维细胞。经过反向蚀斑筛选,获得表达NDV F基因的r DEV-US10-F。对重组区域US10基因测序结果表明,F基因正确插入至US10基因编码区内。将重组病毒传至15代,F基因仍能够稳定表达,表明重组病毒具有稳定的遗传特性,并且与亲本病毒株具有相似的生长曲线。本研究为研发NDV-DEV活载体疫苗奠定了基础。更多还原

【Abstract】 In order to construct the recombinant duck enteritis virus(DEV) expressing F gene of Newcastle disease virus(NDV),the F gene of NDV was amplified by PCR to construct the p US10-TPA-F transfer recombinant plasmid.Then,the p US10-TPA-F and the genomic DNA of recombinant DEV expressing GFP(r DEV-US10-EGFP),were co-transfected into chicken embryo fibroblast(CEF) to generate recombinant virus expressing F protein(r DEV-US10-F),and the r DEV-US10-F was rescued by screening the non-GFP expressing viral plaques.The sequencing result showed that F gene was inserted into the US10 gene of DEV.The F gene was stably expressed from r DEV-US10-F when the recombinant virus was even passed to 15 passages in CEF.In addition,the recombinant virus and the parental virus had similar growth curves.This study would facilitate for developing a Newcastle disease-duck enteritis bivalent live vaccine.更多还原