Autographa californica nucleopolyhedrovirus (AcMNPV) has a large doublestrand DNA genome of approximately 134 kbp and harbors 156 open reading frames (ORFs). To elucidate DNA replication cascade of AcMNPV, we developed a novel baculovirus genome that can be maintained in Escherichia coli as a plasmid and can infect susceptible lepidopteran insect cells. This genome, named bAc-MK, contains a mini-F replicon and a kanamycin resistance marker. Using a convenient Tn7 transposon-based system, pPCS-S, 55 single ORF-truncated mutants were generated by random insertion into bAc-MK genome. These single ORF-truncated mutants were independently transfected into Sf9 cells, 16 of them were found affecting viral replication since they defected in producing polyhedra. Furthermore, to verify the pathogenicity of the single ORF-truncated mutants, the remaining 39 mutants were subjected to bioassay to Spodoptera exigua 3rd instar larvae. Among them, ac9-, ac49-, ac103- and ac105-knockout mutants showed higher mortality compared to that of bAc-MK. These results suggested that these ORFs could be related to pathogenicity of AcMNPV.

• Xue Ying Tao(Department of Agricultural Biotechnology, Seoul National University)
• Jong Yul Roh(Department of Agricultural Biotechnology, Seoul National University)
• Jae Young Choi(Research Institute for Agriculture and Life Sciences, Seoul National University)
• Yong Wang(Department of Agricultural Biotechnology, Seoul National University)
• Qin Liu(Department of Agricultural Biotechnology, Seoul National University)
• Jong Bin Park(Department of Agricultural Biotechnology, Seoul National University)
• Soo Dong Woo(College of Agriculture, Life and Environment Sciences, Chungbuk National University)
• Byung Rae Jin(College of Natural Resources and Life Science, Dong-A University)
• Yeon Ho Je(Department of Agricultural Biotechnology, Seoul National University)