title: Site-specific genetic engineering of the Anopheles gambiae Y chromosome.
abstract: Despite its function in sex determination and its role in driving genome evolution, the Y chromosome remains poorly understood in most species. Y chromosomes are gene-poor, repeat-rich and largely heterochromatic and therefore represent a difficult target for genetic engineering. The Y chromosome of the human malaria vector Anopheles gambiae appears to be involved in sex determination although very little is known about both its structure and function. Here, we characterize a transgenic strain of this mosquito species, obtained by transposon-mediated integration of a transgene construct onto the Y chromosome. Using meganuclease-induced homologous repair we introduce a site-specific recombination signal onto the Y chromosome and show that the resulting docking line can be used for secondary integration. To demonstrate its utility, we study the activity of a germ-line-specific promoter when located on the Y chromosome. We also show that Y-linked fluorescent transgenes allow automated sex separation of this important vector species, providing the means to generate large single-sex populations. Our findings will aid studies of sex chromosome function and enable the development of male-exclusive genetic traits for vector control.
authors.affiliation: Department of Life Sciences, South Kensington Campus, Imperial College London, London SW7 2AZ, United Kingdom;
authors.forename: Federica
authors.initials: F
authors.lastname: Bernardini
authors.affiliation: Department of Life Sciences, South Kensington Campus, Imperial College London, London SW7 2AZ, United Kingdom;
authors.forename: Roberto
authors.initials: R
authors.lastname: Galizi
authors.affiliation: Department of Life Sciences, South Kensington Campus, Imperial College London, London SW7 2AZ, United Kingdom;
authors.forename: Miriam
authors.initials: M
authors.lastname: Menichelli
authors.affiliation: Dipartimento di Medicina Sperimentale Via Gambuli, Centro di Genomica Funzionale, University of Perugia, 06132 Perugia, Italy; and.
authors.forename: Philippos-Aris
authors.initials: PA
authors.lastname: Papathanos
authors.affiliation: Dipartimento di Medicina Sperimentale Via Gambuli, Centro di Genomica Funzionale, University of Perugia, 06132 Perugia, Italy; and.
authors.forename: Vicky
authors.initials: V
authors.lastname: Dritsou
authors.affiliation: Institut de Biologie Moléculaire et Cellulaire, Institut National de la Santé et de la Recherche Médicale U963, Centre National de la Recherche Scientifique, Unité Propre de Recherche 9022, 67084 Strasbourg, France.
authors.forename: Eric
authors.initials: E
authors.lastname: Marois
authors.affiliation: Department of Life Sciences, South Kensington Campus, Imperial College London, London SW7 2AZ, United Kingdom;Dipartimento di Medicina Sperimentale Via Gambuli, Centro di Genomica Funzionale, University of Perugia, 06132 Perugia, Italy; and.
authors.forename: Andrea
authors.initials: A
authors.lastname: Crisanti
authors.affiliation: Department of Life Sciences, South Kensington Campus, Imperial College London, London SW7 2AZ, United Kingdom; n.windbichler@imperial.ac.uk.
authors.forename: Nikolai
authors.initials: N
authors.lastname: Windbichler
mesh_terms: D000818:Animals; D000852:Anopheles / Q000235:genetics*; D001483:Base Sequence; D022202:Chromosomes, Artificial, Bacterial; D059006:Chromosomes, Insect / Q000235:genetics*; D017931:DNA Primers / Q000235:genetics; D005434:Flow Cytometry; D005453:Fluorescence; D055879:Gene Knock-In Techniques; D018014:Gene Transfer Techniques; D005818:Genetic Engineering / Q000379:methods*; D017404:In Situ Hybridization, Fluorescence; D008297:Male; D008969:Molecular Sequence Data; D017423:Sequence Analysis, RNA; D013091:Spermatogenesis / Q000502:physiology; D019076:Transgenes / Q000235:genetics; D014998:Y Chromosome / Q000235:genetics*
chemical_list: D017931:DNA Primers
keywords: SIT; biotechnology