Expression of Bombyx mori nucleopolyhedrovirus ORF76 in permissive and non-permissive cell lines by a novel Bac-to-Bac/BmNPV baculovirus expression system.
Open reading frame 76 of Bombyx mori nucleopolyhedrovirus (BmNPV), designated as Bm76, is a gene whose function is completely unknown. With EGFP fused to the 3' terminal of Bm76 as the reporter gene and BmNPV bacmid as the expression vector, a recombinant bacmid was successfully constructed expressing Bm76-EGFP fusion protein under the control of polyhedrin promoter in Bombyx mori cells (Bm cells), BmNPV's permissive cell line, laying the foundation for rescue experiment of Bm76 deletion mutant. Moreover, the supernatant from Bm cells transfected with the recombinant bacmid was used to infect Trichoplusia Ni cells (Tn cells), BmNPV's non-permissive cell line. Unexpectedly, the expression of Bm76-EGFP fusion protein in some Tn cells was detected, implying that viral DNA was replicated in these cells. The causes are being studied for the inability of BmNPV to produce enough viable budded viruses in Tn cells despite of viral DNA replication.
Su
Wu-Jie
WJ
Institute of Life Sciences, JiangSu University, Xuefu Road 301, Zhenjiang 212013, PR China.
Wu
Yan
Y
Wu
Hui-Ling
HL
Zhu
Shan-Ying
SY
Wang
Wen-Bing
WB
D000818:Animals; D012831:Bombyx / Q000235:genetics / Q000378:metabolism / Q000821:virology*; D002460:Cell Line; D022202:Chromosomes, Artificial, Bacterial; D004274:DNA, Recombinant / Q000235:genetics / Q000378:metabolism; D004279:DNA, Viral / Q000235:genetics* / Q000378:metabolism*; D017930:Genes, Reporter; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins; D017924:Nucleopolyhedroviruses / Q000502:physiology*; D000077183:Occlusion Body Matrix Proteins; D016366:Open Reading Frames; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000235:genetics* / Q000378:metabolism*; D015678:Viral Structural Proteins / Q000235:genetics; D014779:Virus Replication
D004274:DNA, Recombinant; D004279:DNA, Viral; D000077183:Occlusion Body Matrix Proteins; D011993:Recombinant Fusion Proteins; D015678:Viral Structural Proteins; C485184:enhanced green fluorescent protein; C009829:polyhedrin protein, Nucleopolyhedrovirus; D049452:Green Fluorescent Proteins