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Bacteriophage phiC31 produces the enzyme integrase that allows the insertion of the phage genome into its bacterial host. This enzyme recognizes a specific DNA sequence in the phage (attP) and a different sequence in the bacterium (attB). Recombination between these sites leads to integration in a reaction that requires no accessory factors. Seminal studies by the Calos laboratory demonstrated that the phiC31 integrase was capable of integrating plasmid with an attB site into mammalian genomes at sites that approximated the attP site. We describe the use of attB-containing plasmids with insulated reporter genes for the successful integration of DNA into Xenopus embryos. The method offers a way to produce transgenic embryos without manipulation of sperm nuclei using microinjection methods that are standard for experiments in Xenopus laevis. The method aims to allow the non-mosaic controlled expression of new genetic material in the injected embryo and compares favorably with the time that is normally taken to analyze embryos injected with mRNAs, plasmids, morpholinos or oligonucleotides.
Allen,
Transgenic Xenopus laevis embryos can be generated using phiC31 integrase.
2005, Pubmed,
Xenbase
Allen,
Transgenic Xenopus laevis embryos can be generated using phiC31 integrase.
2005,
Pubmed
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Xenbase Bell,
The protein CTCF is required for the enhancer blocking activity of vertebrate insulators.
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Pubmed Bertoni,
Enhancement of plasmid-mediated gene therapy for muscular dystrophy by directed plasmid integration.
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Pubmed Chalberg,
Integration specificity of phage phiC31 integrase in the human genome.
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Pubmed Chalberg,
phiC31 integrase confers genomic integration and long-term transgene expression in rat retina.
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Pubmed Ginsburg,
Site-specific integration with phiC31 integrase for prolonged expression of therapeutic genes.
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Pubmed Groth,
Construction of transgenic Drosophila by using the site-specific integrase from phage phiC31.
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Pubmed Groth,
A phage integrase directs efficient site-specific integration in human cells.
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Pubmed Held,
In vivo correction of murine hereditary tyrosinemia type I by phiC31 integrase-mediated gene delivery.
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Pubmed Hollis,
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Xenbase Olivares,
Site-specific genomic integration produces therapeutic Factor IX levels in mice.
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Pubmed Thomason,
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Pubmed Thyagarajan,
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Pubmed Thyagarajan,
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Pubmed Thyagarajan,
Site-specific genomic integration in mammalian cells mediated by phage phiC31 integrase.
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Pubmed West,
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Pubmed
