title: Identification of rat Rosa26 locus enables generation of knock-in rat lines ubiquitously expressing tdTomato.
abstract: Recent discovery of a method for derivation and culture of germline-competent rat pluripotent stem cells (PSCs) enables generation of transgenic rats or knock-out rats via genetic modification of such PSCs. This opens the way to use rats, as is routine in mice, for analyses of gene functions or physiological features. In mouse or human, one widely used technique to express a gene of interest stably and ubiquitously is to insert that gene into the Rosa26 locus via gene targeting of PSCs. Rosa26 knock-in mice conditionally expressing a reporter or a toxin gene have contributed to tracing or ablation of specific cell lineages. We successfully identified a rat orthologue of the mouse Rosa26 locus. Insertion of tdTomato, a variant of red fluorescent protein, into the Rosa26 locus of PSCs of various rat strains allows ubiquitous expression of tdTomato. Through germline transmission of one Rosa26-tdTomato knock-in embryonic stem cell line, we also obtained tdTomato knock-in rats. These expressed tdTomato ubiquitously throughout their bodies, which indicates that the rat Rosa26 locus conserves functions of its orthologues in mouse and human. The new tools described here (targeting vectors, knock-in PSCs, and rats) should be useful for a variety of research using rats.
authors.affiliation: Division of Stem Cell Therapy, Center for Stem Cell Biology and Regenerative Medicine, Institute of Medical Science, University of Tokyo, Tokyo, Japan.
authors.forename: Toshihiro
authors.initials: T
authors.lastname: Kobayashi
authors.forename: Megumi
authors.initials: M
authors.lastname: Kato-Itoh
authors.forename: Tomoyuki
authors.initials: T
authors.lastname: Yamaguchi
authors.forename: Chihiro
authors.initials: C
authors.lastname: Tamura
authors.forename: Makoto
authors.initials: M
authors.lastname: Sanbo
authors.forename: Masumi
authors.initials: M
authors.lastname: Hirabayashi
authors.forename: Hiromitsu
authors.initials: H
authors.lastname: Nakauchi
mesh_terms: D000818:Animals; D001755:Blastocyst / Q000166:cytology / Q000378:metabolism; D053595:Embryonic Stem Cells / Q000166:cytology / Q000378:metabolism; D055879:Gene Knock-In Techniques / Q000379:methods*; D018390:Gene Targeting; D056426:Genetic Loci / Q000235:genetics*; D006801:Humans; D008164:Luminescent Proteins / Q000378:metabolism*; D051379:Mice; D039904:Pluripotent Stem Cells / Q000166:cytology / Q000378:metabolism; D051381:Rats; D055647:Rats, Transgenic; D000097573:Red Fluorescent Protein
chemical_list: D008164:Luminescent Proteins; D000097573:Red Fluorescent Protein