Epstein-Barr virus plasmid model system for analyzing recombination in human cells.
Homologous recombination stimulated by a double-strand break at a desired target site offers a method to achieve site-specific integration useful for gene therapy and other genetic engineering. To test parameters needed for this strategy, we developed an Epstein-Barr virus shuttle vector model system as a genetic tool. This extrachromosomal plasmid assay system has several advantages over a chromosomal assay. The system detects all classes of recombination events without selection and allows rapid analysis of the frequency and nature of recombination events. We found that a double-strand break at the target site stimulated a large increase in recombination frequency. The resulting recombinants included one-sided insertion events, as well as two-sided or gene conversion events. A circular donor substrate was more effective in recombination than linearized donor DNA.
Phillips
J E
JE
Department of Genetics, Stanford University School of Medicine, Stanford, California 94305-5120, USA.
Thyagarajan
B
B
Calos
M P
MP
D001483:Base Sequence; D002460:Cell Line; D005822:Genetic Vectors; D004854:Herpesvirus 4, Human / Q000235:genetics*; D006801:Humans; D007763:Lac Operon; D008957:Models, Genetic; D016254:Mutagenesis, Insertional; D010957:Plasmids / Q000235:genetics*; D011995:Recombination, Genetic