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- Title
Characterization of AcMNPV with a deletion of me53 gene.
- Authors
Jinwen Wang; Riqiang Deng; Xunzhang Wang
- Abstract
Abstract??TheAutographa Californicamultiple nucleopolyhedrovirus (AcMNPV)me53gene, which was previously reported as one of the major early-transcribed genes, was deleted through homologous recombination from an AcMNPV genome propagated as a bacmid DNA inE. coli, generating ame53gene knockout bacmid. Green fluorescent protein (GFP) expression analysis and supernatant passage assay revealed that theme53 knockout bacmid was unable to replicate in cell culture, whileme53repair bacmid, which was generated by reinsertion of theme53 gene into the mutant at a different locus (the gentamicin locus) with ET-recombination technique, resumed viral replication ability at wild-type levels, indicating that the defective phenotype of theme53knockout virus was directly due to the deletion of theme53-ORF. Subsequent electron microscopy revealed that theme53 knockout bacmid failed to form nucleocapsid in the nuclei of the transfected cells, though viral infection seemed to be initiated. Meanwhile, real-time PCR analysis based on SYBR Green fluorescence indicated abolishment of the viral DNA replication byme53gene inactivation. Thus, it is demonstrated for the first time thatme53 knockout blocked viral DNA replication, nucleocapsid formation, and consequent BV and ODV production.
- Subjects
GENES; DEOXYRIBOSE; DNA; NUCLEIC acids
- Publication
Virus Genes, 2007, Vol 34, Issue 2, p223
- ISSN
0920-8569
- Publication type
Article