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A novel method using baculovirus-mediated gene transfer for production of recombinant adeno-associated virus vectors

Center for Genomics Research, Karolinska Institute, SE-17177 Stockholm, Sweden¹
CNRS UMR 1582, Vectorologie et Transfert de Génes, Institut Gustave Roussy, 94805 Villejuif Cedex, France²

Author for correspondence: Kerstin Sollerbrant. Mailing address: Ludwig Institute for Cancer Research, Stockholm Branch, Box 240, SE-17177 Stockholm, Sweden. Fax +46 8 332812. e-mail kerstin.sollerbrant{at}licr.ki.se

The baculovirus Autographa californica multiple nucleopolyhedrosis virus causes non-productive infection in mammalian cells. Recombinant baculovirus therefore has the capability to transfer and express heterologous genes in these cells if a mammalian promoter governs the gene of interest. We have investigated the possibility of using baculovirus as a tool to produce recombinant adeno-associated virus (rAAV). AAV has become increasingly popular as a vector for gene therapy and functional genomics efforts, although its use is hampered by the lack of a simple and efficient vector production method. We show here that co-infection of mammalian producer cells with three viruses – a baculovirus containing the reporter gene flanked by AAV ITRs, a baculovirus expressing the AAV rep gene and a helper adenovirus expressing the AAV cap gene – produces infectious rAAV particles. This baculovirus-based chimeric vector method may in future improve large-scale rAAV vector preparations and circumvent present-day problems associated with rAAV production.

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