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.
This is a preview of subscription content, access via your institution
Access options
Subscribe to this journal
Receive 12 print issues and online access
$259.00 per year
only $21.58 per issue
Buy this article
- Purchase on Springer Link
- Instant access to full article PDF
Prices may be subject to local taxes which are calculated during checkout
Similar content being viewed by others
Author information
Authors and Affiliations
Rights 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
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1038/sj.gt.3300680
Keywords
This article is cited by
-
Recent advances in various adeno-associated viruses (AAVs) as gene therapy agents in hepatocellular carcinoma
Virology Journal (2024)
-
Comparison of three AUC techniques for the determination of the loading status and capsid titer of AAVs
European Biophysics Journal (2023)
-
Focal loss of the paranodal domain protein Neurofascin155 in the internal capsule impairs cortically induced muscle activity in vivo
Molecular Brain (2020)
-
Antigen-selective modulation of AAV immunogenicity with tolerogenic rapamycin nanoparticles enables successful vector re-administration
Nature Communications (2018)
-
Molecular design for recombinant adeno-associated virus (rAAV) vector production
Applied Microbiology and Biotechnology (2018)