Herpes simplex virus-induced “Rolling circle” amplification of SV40 DNA sequences in a transformed hamster cell line correlates with tandem integration of the SV40 genome

ABSTRACTAdeno-associated virus (AAV) genome replication only occurs in the presence of a co-infecting helper virus such as adenovirus type 5 (AdV5) or herpes simplex virus type 1 (HSV-1). AdV5-supported replication of the AAV genome has been described to occur in a strand-displacement rolling hairpin mechanism initiated at the AAV 3’ inverted terminal repeat (ITR) end. It has been assumed that the same mechanism applies to HSV-1-supported AAV genome replication. We demonstrate the formation of double-stranded head-to-tail concatemers of AAV genomes in presence of HSV-1, and thus provide evidence for an unequivocal rolling circle amplification (RCA) mechanism. This study reveals the ability of AAV to modify the canonical rolling hairpin replication mechanism and to mimic the replication strategy of a co-infecting herpesvirus. This stands in contrast to the textbook model of AAV genome replication when HSV-1 is the helper virus. Furthermore, we introduce nanopore sequencing as a novel, high-throughput approach to study viral genome replication in unprecedented detail.

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The DNA Sequences of the Long Repeat Region and Adjoining Parts of the Long Unique Region in the Genome of Herpes Simplex Virus Type 1

Journal of General Virology ◽

10.1099/0022-1317-69-11-2831 ◽

1988 ◽

Vol 69 (11) ◽

pp. 2831-2846 ◽

Cited By ~ 70

Author(s):

L. J. Perry ◽

D. J. McGeoch

Keyword(s):

Herpes Simplex Virus ◽

Herpes Simplex ◽

Virus Type ◽

Dna Sequences ◽

Herpes Simplex Virus Type ◽

Repeat Region ◽

Unique Region ◽

Simplex Virus

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Adaptation of a retrovirus as a eucaryotic vector transmitting the herpes simplex virus thymidine kinase gene.

Molecular and Cellular Biology ◽

10.1128/mcb.2.4.426 ◽

1982 ◽

Vol 2 (4) ◽

pp. 426-436 ◽

Cited By ~ 89

Author(s):

C J Tabin ◽

J W Hoffmann ◽

S P Goff ◽

R A Weinberg

Keyword(s):

Herpes Simplex Virus ◽

Herpes Simplex ◽

Thymidine Kinase ◽

Dna Sequences ◽

Leukemia Virus ◽

Chimeric Virus ◽

Animal Cells ◽

Kinase Gene ◽

General Applicability ◽

Simplex Virus

We investigated the feasibility of using retroviruses as vectors for transferring DNA sequences into animal cells. The thymidine kinase (tk) gene of herpes simplex virus was chosen as a convenient model. The internal BamHI fragments of a DNA clone of Moloney leukemia virus (MLV) were replaced with a purified BamHI DNA segment containing the tk gene. Chimeric genomes were created carrying the tk insert in both orientations relative to the MLV sequence. Each was transfected into TK- cells along with MLV helper virus, and TK+ colonies were obtained by selection in the presence of hypoxanthine, aminopterin, and thymidine (HAT). Virus collected from TK+-transformed, MLV producer cells passed the TK+ phenotype to TK- cells. Nonproducer cells were isolated, and TK+ transducing virus was subsequently rescued from them. The chimeric virus showed single-hit kinetics in infections. Virion and cellular RNA and cellular DNA from infected cells were all shown to contain sequences which hybridized to both MLV- and tk-specific probes. The sizes of these sequences were consistent with those predicted for the chimeric virus. In all respects studied, the chimeric MLV-tk virus behaved like known replication-defective retroviruses. These experiments suggest great general applicability of retroviruses as eucaryotic vectors.

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