(NAS Colloquium) Genetic Engineering of Viruses and Viral Vectors (1996) / Chapter Skim
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Site-specific integration by adeno-associated virus
Site-specific integration by adeno-associated virus
Pages 11288-11294
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From page 11288... ...
Use of a functional model system for AAV DNA integration into AAVS1 has allowed us to conclude that the recombination event is directed by cellular DNA sequences. Recombinant junctions isolated from our integration assay were analyzed and showed characteristics similar to those found in latently infected cell lines.
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From page 11289... ...
In the presence of helper virus (i.e., the permissive state) Rep 68/78 is required for AAV gene expression and transactivates expression of the structural proteins; it is also required for viral DNA replication and rescue of the integrated viral genome.
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From page 11290... ...
Nucleotide numbers indicated are given with respect toAAVSl. M26, enhancer of meiotic gene conversion in fission yeast; CRE, cyclic AMP response element; Spl, transcription factor Spl consensus sequence; TRS, terminal resolution site; RBS, Rep binding site.
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From page 11291... ...
In most cases the AAV sequences missing were from the part of the AAV genome encoding the REP genes. The integrated DNA sequence extended from a point within the internal AAV sequence through the ITR at the 3' end of the genome, which again had been elongated to form a head-to-tail junction and then extended in another sequence to map position 5 (as in the two full-length inserts described above)
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From page 11292... ...
(~ After synthesis of AAV DNA sequences a third template strand switch back onto AAVS1 results in a second link between viral and host DNA sequences.
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From page 11293... ...
Use of uninfected cell extract leads to premature strand switching and the consequent interruption of the normal replication process with the synthesis of defective DNA molecules (35~. We believe that the enhanced probability of strand switching during DNA synthesis in the absence of a helper virus coinfection plays a major role in the integration process.
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From page 11294... ...
11294 Colloquium Paper: Linden et al.
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