Open AccessCloning of adeno-associated virus into pBR322: rescue of intact virus from the recombinant plasmid in human cells.
Author(s) -
R. Jude Samulski,
Kenneth I. Berns,
Ming Tan,
Nicholas Muzyczka
Publication year - 1982
Publication title -
proceedings of the national academy of sciences of the united states of america
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.011
H-Index - 771
eISSN - 1091-6490
pISSN - 0027-8424
DOI - 10.1073/pnas.79.6.2077
Subject(s) - plasmid , recombinant dna , pbr322 , adeno associated virus , transfection , biology , virology , microbiology and biotechnology , virus , dna , t dna binary system , plasmid preparation , cloning (programming) , molecular cloning , gene , vector (molecular biology) , genetics , complementary dna , computer science , programming language
We have cloned intact duplex adeno-associated virus (AAV) DNA into the bacterial plasmid pBR322. The AAV genome could be rescued from the recombinant plasmid by transfection of the plasmid DNA into human cells with adenovirus 5 as helper. The efficiency of rescue from the plasmid was sufficiently high to produce yields of AAV DNA comparable to those observed after transfection with equal amounts of purified virion DNA. Thus, the recombinant plasmid itself may be a model for studying the rescue of a latent AAV viral infection. In addition, the efficient rescue of viable AAV from the recombinant plasmid should facilitate the genetic analysis of AAV. Finally, the results of an analysis of the DNA from rescued virions indicate that an inversion of the AAV terminal sequences occurred during replication.