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ABSTRACT The purpose of this work was to develop an efficient method for the production of adeno-associated virus (AAV) vectors in the absence of helper virus. The adenovirus regions that

mediate AAV vector replication were identified and assembled into a helper plasmid. These included the VA, E2A and E4 regions. When this helper plasmid was cotransfected into 293 cells,

along with plasmids encoding the AAV vector, and rep and cap genes, AAV vector was produced as efficiently as when using adenovirus infection as a source of help. CMV-driven constructs

expressing the E4orf6 and the 72-Mr, E2A proteins were able to functionally replace the E4 and E2A regions, respectively. Therefore the minimum set of genes required to produce AAV helper

activity equivalent to that provided by adenovirus infection consists of, or is a subset of, the following genes: the E4orf6 gene, the 72-Mr, E2A protein gene, the VA RNA genes and the E1

region. AAV vector preparations made with adenovirus and by the helper virus-free method were essentially indistinguishable with respect to particle density, particle to infectivity ratio,

capsimer ratio and efficiency of muscle transduction in vivo. Only AAV vector preparations made by the helper virus-free method were not reactive with anti-adenovirus sera. Access through

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RNA VIRUSES Article Open access 08 June 2021 AUTHOR INFORMATION AUTHORS AND AFFILIATIONS * Avigen, Inc., Alameda, CA, USA T Matsushita, S Elliger, C Elliger, G Podsakoff, GJ Kurtzman & P

Colosi * University of Southern California School of Medicine, Los Angeles, CA, USA T Matsushita * Department of Molecular Biology and Biochemistry, University of California, Irvine, CA,

USA L Villarreal * Department of Urology, University of Southern California School of Medicine, Los Angeles, CA, USA Y Iwaki * Childrens Hospital Oakland Research Institute, 747 52nd Street,

Oakland, 95609, CA S Elliger * California Academy of Sciences, Golden Gate Park, San Francisco, 94118, CA, USA C Elliger Authors * T Matsushita View author publications You can also search

for this author inPubMed Google Scholar * S Elliger View author publications You can also search for this author inPubMed Google Scholar * C Elliger View author publications You can also

search for this author inPubMed Google Scholar * G Podsakoff View author publications You can also search for this author inPubMed Google Scholar * L Villarreal View author publications You

can also search for this author inPubMed Google Scholar * GJ Kurtzman View author publications You can also search for this author inPubMed Google Scholar * Y Iwaki View author publications

You can also search for this author inPubMed Google Scholar * P Colosi View author publications You can also search for this author inPubMed Google Scholar RIGHTS AND PERMISSIONS Reprints

and permissions ABOUT THIS ARTICLE CITE THIS ARTICLE Matsushita, T., Elliger, S., Elliger, C. _et al._ Adeno-associated virus vectors can be efficiently produced without helper virus. _Gene

Ther_ 5, 938–945 (1998). https://doi.org/10.1038/sj.gt.3300680 Download citation * Received: 08 January 1998 * Accepted: 10 February 1998 * Published: 10 July 1998 * Issue Date: 01 July 1998

* DOI: https://doi.org/10.1038/sj.gt.3300680 SHARE THIS ARTICLE Anyone you share the following link with will be able to read this content: Get shareable link Sorry, a shareable link is not

currently available for this article. Copy to clipboard Provided by the Springer Nature SharedIt content-sharing initiative KEYWORDS * AAV vector production * AAV helper genes * adenovirus