title: Efficient mouse transgenesis using Gateway-compatible ROSA26 locus targeting vectors and F1 hybrid ES cells.
abstract: The ability to rapidly and efficiently generate reliable Cre/loxP conditional transgenic mice would greatly complement global high-throughput gene targeting initiatives aimed at identifying gene function in the mouse. We report here the generation of Cre/loxP conditional ROSA26-targeted ES cells within 3-4 weeks by using Gateway cloning to build the target vectors. The cDNA of the gene of interest can be expressed either directly by the ROSA26 promoter providing a moderate level of expression or by a CAGG promoter placed in the ROSA26 locus providing higher transgene expression. Utilization of F1 hybrid ES cells with exceptional developmental potential allows the production of germ line transmitting, fully or highly ES cell-derived mice by aggregation of cells with diploid embryos. The presented streamlined procedures accelerate the examination of phenotypical consequences of transgene expression. It also provides a unique tool for comparing the biological activity of polymorphic or splice variants of a gene, or products of different genes functioning in the same or parallel pathways in an overlapping manner.
authors.affiliation: Department for Molecular Biomedical Research, VIB, Gent, Belgium.
authors.forename: Omar
authors.initials: O
authors.lastname: Nyabi
authors.forename: Michael
authors.initials: M
authors.lastname: Naessens
authors.forename: Katharina
authors.initials: K
authors.lastname: Haigh
authors.forename: Agnieszka
authors.initials: A
authors.lastname: Gembarska
authors.forename: Steven
authors.initials: S
authors.lastname: Goossens
authors.forename: Marion
authors.initials: M
authors.lastname: Maetens
authors.forename: Sarah
authors.initials: S
authors.lastname: De Clercq
authors.forename: Benjamin
authors.initials: B
authors.lastname: Drogat
authors.forename: Lieven
authors.initials: L
authors.lastname: Haenebalcke
authors.forename: Sonia
authors.initials: S
authors.lastname: Bartunkova
authors.forename: Ilse
authors.initials: I
authors.lastname: De Vos
authors.forename: Bram
authors.initials: B
authors.lastname: De Craene
authors.forename: Mansour
authors.initials: M
authors.lastname: Karimi
authors.forename: Geert
authors.initials: G
authors.lastname: Berx
authors.forename: Andras
authors.initials: A
authors.lastname: Nagy
authors.forename: Pierre
authors.initials: P
authors.lastname: Hilson
authors.forename: Jean-Christophe
authors.initials: JC
authors.lastname: Marine
authors.forename: Jody J
authors.initials: JJ
authors.lastname: Haigh
mesh_terms: D000483:Alleles; D000818:Animals; D002460:Cell Line; D003001:Cloning, Molecular / Q000379:methods*; D004171:Diploidy; D053595:Embryonic Stem Cells / Q000378:metabolism*; D018390:Gene Targeting / Q000379:methods*; D005822:Genetic Vectors; D006822:Hybrid Cells; D051379:Mice; D008822:Mice, Transgenic; D011506:Proteins / Q000235:genetics*; D022661:RNA, Untranslated; D011995:Recombination, Genetic; D019076:Transgenes
chemical_list: C110853:Gt(ROSA)26Sor non-coding RNA, mouse; D011506:Proteins; D022661:RNA, Untranslated