title: Recombinase-mediated mouse transgenesis by intracytoplasmic sperm injection.
abstract: The low efficiency of current microinjection-based animal transgenesis techniques is largely the result of poor embryo survival. We have developed a new, bacterial recombinase-based transgenesis method. Intracytoplasmic sperm injection (ICSI) of single stranded DNA (ssDNA) complexed with E. coli recombinase RecA into mouse metaphaseII (MII) arrested oocytes resulted in RecA-dependent transgenesis. This approach offers significant advantages over pronuclear microinjection and previous ICSI-based transgenesis approaches in terms of improved embryo survival, which translates into greater transgenesis efficiency. It also opens the possibility to attempt experiments, which may affect gene targeting by homologous recombination into DNA of mammalian single celled pre-implantation embryos.
authors.affiliation: Department of Anatomy and Reproductive Biology, University of Hawaii School of Medicine, HI 96822, USA.
authors.forename: Takehito
authors.initials: T
authors.lastname: Kaneko
authors.forename: Stefan
authors.initials: S
authors.lastname: Moisyadi
authors.forename: Ryota
authors.initials: R
authors.lastname: Suganuma
authors.forename: Barbara
authors.initials: B
authors.lastname: Hohn
authors.forename: Ryuzo
authors.initials: R
authors.lastname: Yanagimachi
authors.forename: Pawel
authors.initials: P
authors.lastname: Pelczar
mesh_terms: D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D002467:Cell Nucleus; D004277:DNA, Single-Stranded / Q000008:administration & dosage / Q000378:metabolism; D046149:Embryo Culture Techniques; D004624:Embryo Transfer; D004926:Escherichia coli / Q000201:enzymology; D015870:Gene Expression; D018014:Gene Transfer Techniques; D049452:Green Fluorescent Proteins / Q000235:genetics; D051379:Mice; D008822:Mice, Transgenic; D008845:Microinjections; D045522:Recombinases / Q000008:administration & dosage* / Q000378:metabolism; D020554:Sperm Injections, Intracytoplasmic; D014162:Transfection / Q000379:methods
chemical_list: D004277:DNA, Single-Stranded; D045522:Recombinases; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D000255:Adenosine Triphosphate