title: Application of laser-assisted zona drilling to in vitro fertilization of cryopreserved mouse oocytes with spermatozoa from a subfertile transgenic mouse.
abstract: Development of assisted reproductive technologies is necessary to obtain fertilized oocytes in a subfertile transgenic mouse strain. Here, we showed the application of laser-assisted drilling of the zona pellucida to in vitro fertilization of cryopreserved mouse oocytes with sperm from subfertile transgenic mice (C57BL/6N-Tg(UCP/FAD2)U8 strain). After cryopreservation by vitrification, the recovery and survival rates of the zona-drilled mouse oocytes were 97% (97/100) and 94% (91/97), respectively. In vitro fertilization of the cryopreserved zona-drilled mouse oocytes with sperm from the subfertile transgenic mice was greatly facilitated (60%, 55/91) compared to that of the cryopreserved zona-intact mouse oocytes (11%, 81/768). In vitro fertilized embryos that developed to the 2-cell stage were again cryopreserved by vitrification, and after warming they were transferred into recipient females. Subsequently, six viable offspring were delivered, and all were confirmed to be transgenic mice. These results indicate that laser-assisted zona drilling of oocytes combined with cryopreservation by vitrification may be a useful approach for large-scale production of in vitro fertilized embryos for managing transgenic mouse strains with reproductive disabilities such as subfertile sperm.
authors.affiliation: Institute of Advanced Technology, Kinki University, Wakayama, Japan. anzai@waka.kindai.ac.jp
authors.forename: Masayuki
authors.initials: M
authors.lastname: Anzai
authors.forename: Megumi
authors.initials: M
authors.lastname: Nishiwaki
authors.forename: Miho
authors.initials: M
authors.lastname: Yanagi
authors.forename: Tatsuyuki
authors.initials: T
authors.lastname: Nakashima
authors.forename: Takehito
authors.initials: T
authors.lastname: Kaneko
authors.forename: Yoshitomo
authors.initials: Y
authors.lastname: Taguchi
authors.forename: Mikiko
authors.initials: M
authors.lastname: Tokoro
authors.forename: Seung-Wook
authors.initials: SW
authors.lastname: Shin
authors.forename: Tasuku
authors.initials: T
authors.lastname: Mitani
authors.forename: Hiromi
authors.initials: H
authors.lastname: Kato
authors.forename: Kazuya
authors.initials: K
authors.lastname: Matsumoto
authors.forename: Naomi
authors.initials: N
authors.lastname: Nakagata
authors.forename: Akira
authors.initials: A
authors.lastname: Iritani
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D015925:Cryopreservation; D005260:Female; D005298:Fertility; D005307:Fertilization in Vitro / Q000379:methods*; D007834:Lasers; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D009865:Oocytes; D013094:Spermatozoa; D015044:Zona Pellucida