PMID: 9525651Apr 3, 1998Paper

In vitro packaging of adeno-associated virus DNA

Journal of Virology
X Zhou, Nicholas Muzyczka

Abstract

We have developed an in vitro procedure for packaging of recombinant adeno-associated virus (AAV). By using AAV replicative-form DNA as the substrate, it is possible to synthesize an infectious AAV particle in vitro that can be used to transfer a marker gene to mammalian cells. The packaging procedure requires the presence of both the AAV Rep and capsid proteins. Two kinds of in vitro products can be formed which facilitate DNA transfer. Both are resistant to heat and have a density in cesium chloride gradients that is indistinguishable from that of the in vivo-synthesized wild-type virus. This indicates that the particles formed have the appropriate protein-to-DNA ratio and a structure that shares the heat resistance of mature AAV particles. The two types of particles can be distinguished by their sensitivity to chloroform and DNase I treatment. The chloroform-resistant product is, by several criteria, an authentic AAV particle. In addition to having the correct density and being resistant to treatment with chloroform, DNase I, and heat, this particle is efficiently synthesized only if the AAV genome contains intact terminal repeats, which are known to be required for AAV packaging. It is also precipitated by a monoclonal anti...Continue Reading

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Citations

Mar 30, 2000·The Journal of Gene Medicine·R O Snyder
Jul 13, 2007·Journal of Virology·Peter WardR Michael Linden
Aug 8, 2014·Frontiers in Immunology·Etiena Basner-Tschakarjan, Federico Mingozzi
Aug 3, 1999·Biological Chemistry·H Büeler
Feb 19, 2000·Gene Therapy·P E Monahan, R J Samulski
Apr 19, 2013·Blood·Federico Mingozzi, Katherine A High
Nov 27, 2015·Expert Review of Ophthalmology·Cristy A Ku, Mark E Pennesi
Apr 19, 2020·Human Gene Therapy·Anna C Maurer, Matthew D Weitzman
Sep 6, 2020·American Journal of Medical Genetics. Part C, Seminars in Medical Genetics·Cristy A Ku, Mark E Pennesi
Feb 3, 2007·Biotechnology and Bioengineering·Marc G AucoinAmine A Kamen
May 21, 2013·Current Protocols in Microbiology·Seemin Seher AhmedLi Zhong
Apr 25, 2000·Journal of Virology·R K Hirata, D W Russell
Jan 10, 2018·Biotechnology Journal·Carolin Schmelas, Dirk Grimm
Apr 15, 2021·Molecular Therapy. Methods & Clinical Development·Kamakshi BankotiDaqing Li
Jun 3, 2021·Pharmaceutics·Maxim A Korneyenkov, Andrey A Zamyatnin
Feb 9, 2012·Molecular Therapy : the Journal of the American Society of Gene Therapy·Casey A MaguireJohan Skog

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