title: In vitro culture and in vitro fertilization techniques for prairie voles (Microtus ochrogaster).
abstract: Prairie vole (Microtus ochrogaster) is a highly social animal and is a commonly used animal model for neuropsychopharmacological and psychiatric studies. To date, only a few reports on the development of transgenic prairie voles which was primarily due to the suboptimal development of assisted reproductive technology (ART) in prairie voles. Limitations in ART further hinder the development of genetically modified prairie voles such as the application of conventional gene targeting technologies using embryonic stem (ES) or induced pluripotent stem (iPS) cells to generate chimeric prairie voles. Moreover, recent advancement in genome-editing tools such as transcription activator-like effector nuclease (TALEN) and clustered regulatory interspaced short palindromic repeat (CRISPR)/Cas technology provide an unprecedented opportunity to create gene targeting animal model and the development of ART in prairie voles is necessary for future development of novel transgenic prairie vole model. We have established efficient method for in vitro embryo culture and sperm cryopreservation with high fertilization rate. In G-1 PLUS and G-2 PLUS sequential culture condition, 81.0% (# of Blastocysts/total n) of one-cell embryos developed to blastocysts. In contrary, no embryos were developed to blastocyst stage in KSOM medium (0/total # of embryos in culture). In vitro fertilization rate using fresh and frozen-thawed sperm was 32.6% and 29.3%, respectively. This is the first report of IVF using cryopreserved prairie vole sperm. We employed mouse IVF methods in prairie voles and optimize culture conditions using human G-1/G-2 PLUS sequential culture method that resulted in high embryonic development rate. The development in vole reproductive technology will facilitate the generation of transgenic voles in the future.
authors.affiliation: Laboratory of Molecular Biology, Department of Molecular and Cell Biology, Graduate School of Agricultural Science, Tohoku University, 1-1 Tsutsumidori-Amamiyamachi, Aoba-ku, Sendai, Miyagi 981-8555, Japan.
authors.forename: Kengo
authors.initials: K
authors.lastname: Horie
authors.affiliation: Laboratory of Molecular Biology, Department of Molecular and Cell Biology, Graduate School of Agricultural Science, Tohoku University, 1-1 Tsutsumidori-Amamiyamachi, Aoba-ku, Sendai, Miyagi 981-8555, Japan.
authors.forename: Shizu
authors.initials: S
authors.lastname: Hidema
authors.affiliation: Department of Obstetrics and Gynecology, Juntendo University, 2-1-1, Hongo, Bunkyo-ku, Tokyo 113-8421, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Hirayama
authors.affiliation: Laboratory of Molecular Biology, Department of Molecular and Cell Biology, Graduate School of Agricultural Science, Tohoku University, 1-1 Tsutsumidori-Amamiyamachi, Aoba-ku, Sendai, Miyagi 981-8555, Japan. Electronic address: knishimori@m.tohoku.ac.jp.
authors.forename: Katsuhiko
authors.initials: K
authors.lastname: Nishimori
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D003411:Arvicolinae / Q000502:physiology*; D005260:Female; D005307:Fertilization in Vitro; D008297:Male
keywords: Assisted reproductive technology (ART); In vitro culture; In vitro fertilization; Prairie vole