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Rescue of the adeno-associated virus genome from a plasmid vector: evidence for rescue by replication.

Artikel i vetenskaplig tidskrift
Författare Peter Ward
Per Elias
R Michael Linden
Publicerad i Journal of virology
Volym 77
Nummer/häfte 21
Sidor 11480-90
ISSN 0022-538X
Publiceringsår 2003
Publicerad vid Institutionen för medicinsk och fysiologisk kemi
Sidor 11480-90
Språk en
Länkar www.ncbi.nlm.nih.gov/entrez/query.f...
Ämnesord DNA Replication, DNA-Binding Proteins, genetics, metabolism, DNA-Directed DNA Polymerase, genetics, metabolism, Dependovirus, genetics, physiology, Genetic Vectors, Genome, Viral, Herpesvirus 1, Human, genetics, metabolism, Humans, Plasmids, genetics, RNA, Viral, Transfection, Viral Proteins, genetics, metabolism, Virus Replication
Ämneskategorier Virologi, Medicinsk bioteknologi (med inriktning mot cellbiologi (inklusive stamcellsbiologi), molekylärbiologi, mikrobiologi, biokemi eller biofarmaci)

Sammanfattning

In cultured cells, adeno-associated virus (AAV) replication requires coinfection with a helper virus, either adenovirus or herpesvirus. In the absence of helper virus coinfection AAV can integrate its genome site specifically into the AAVS1 region of chromosome 19. Upon subsequent infection with a helper virus, the AAV genome is released from chromosome 19 by a process termed rescue, and productive replication ensues. The AAV genome cloned into a plasmid vector can also serve to initiate productive AAV replication. When such constructs are transfected into cells and those cells are simultaneously or subsequently infected with a helper virus, the AAV genome is released from the plasmid. This process is thought to serve as a model for rescue from the human genomic site. In this report we present a model for rescue of AAV genomes by replication. A hallmark of this model is the production of a partially single-stranded and partially double-stranded molecule. We show that the AAV2 Rep 68 protein, together with the UL30/UL42 herpes simplex virus type 1 DNA polymerase and the UL29 single-strand DNA binding protein ICP8, is sufficient to efficiently and precisely rescue AAV from a plasmid in a way that is dependent on the AAV inverted terminal repeat sequence.

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