title: Use of phage φC31 integrase as a tool for zebrafish genome manipulation.
abstract: On the strengths of forward genetics and embryology, the zebrafish Danio rerio has become an ideal system for the study of early vertebrate development. However, additional tools will be needed to perform more sophisticated analyses and to successfully carry this model into new areas of study such as adult physiology, cancer, and aging. As improved tools make transgenesis more and more efficient, the stage has been set for precise modification of the zebrafish genome such as are done in other model organisms. Genome engineering strategies employing site-specific recombinase (SSR) systems such as Cre/lox and Flp/FRT have become invaluable to the study of gene function in the mouse and Drosophila and are now being exploited in zebrafish as well. My laboratory has begun to use another such SSR, the integrase encoded by the Streptomyces bacteriophage PhiC31, for manipulation of the zebrafish genome. The PhiC31 integrase promotes recombination between an attachment site in the phage (attP) and another on the bacterial chromosome (attB). Here I describe strategies using the PhiC31 integrase to mediate recombination of transgenes containing attP and attB sites in cis to excise elements with spatial and temporal specificity. The feasibility of the intramolecular recombination approach having been established, I discuss prospects for employing PhiC31 integrase for intermolecular recombination, i.e., transgene integration at defined sites in the genome.
authors.affiliation: Department of Human and Molecular Genetics and Massey Cancer Center, Virginia Commonwealth University School of Medicine, Box 980033, Richmond, Virginia, USA.
authors.forename: James A
authors.initials: JA
authors.lastname: Lister
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D001435:Bacteriophages / Q000201:enzymology*; D004254:DNA Nucleotidyltransferases / Q000235:genetics / Q000378:metabolism; D005260:Female; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism; D017930:Genes, Reporter; D005822:Genetic Vectors; D016678:Genome; D019426:Integrases / Q000235:genetics* / Q000378:metabolism; D007814:Larva / Q000235:genetics / Q000378:metabolism; D008297:Male; D016254:Mutagenesis, Insertional / Q000379:methods*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014764:Viral Proteins / Q000235:genetics* / Q000378:metabolism; D015027:Zebrafish / Q000235:genetics*
chemical_list: D005656:Fungal Proteins; D011994:Recombinant Proteins; D014764:Viral Proteins; C045073:Cre recombinase; D004254:DNA Nucleotidyltransferases; C045119:FLP recombinase; D019426:Integrases