Replication of rep-cap genes is essential for the high-efficiency production of recombinant AAV

Human Gene Therapy
P D Fan, J Y Dong

Abstract

Adenoassociated virus (AAV) has been developed as a vector for gene transfer because of its advantageous features: it is nonpathogenic, naturally replication-defective; it infects growth-arrested cells, and can transfer the therapeutic gene without co-delivery of any viral genes. However, a major obstacle in conducting systematic studies of AAV-mediated gene transfer in animal models is the difficulty of obtaining large quantities of recombinant virus. Recent development of AAV packaging cell lines has simplified the procedure of producing recombinant AAV (rAAV). However, the efficacy of producing large quantities of rAAV with these cell lines is yet to be demonstrated. In this study we have analyzed the difference between the replication of wild-type AAV and the production of rAAV. Using a combined single-plasmid system that carries both an AAV vector and the rep-cap genes, we have demonstrated that the AAV vector replicates to high number of copies whereas the rep-cap sequences remain unamplified in the virus-producing cells, When the copy number of rep-cap genes was increased by varying the vector/rep-cap ratio in the transfection mixture, the titer of rAAV increased proportionally. Thus, the titer of rAAV is limited by the ...Continue Reading

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Citations

Nov 6, 2003·Journal of Virological Methods·Alistair WhitewayRobert Anderson
Jan 29, 1999·Journal of Virological Methods·J Chirico, J P Trempe
Feb 14, 2002·Kidney International·K Reed Clark
Aug 26, 1998·Proceedings of the National Academy of Sciences of the United States of America·L ZhangJ Y Dong
Jul 13, 2007·Journal of Virology·Peter WardR Michael Linden
Jun 7, 2005·Expert Opinion on Biological Therapy·Luisa BarzonGiorgio Palù
Oct 19, 2005·Gene Therapy·O-W MertenA M Douar
Jul 15, 2000·Journal of Biomedical Science·J Tal

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