2細胞期胚の分離によるマウスー卵性双子の作出

抄録

The present study was examine the viavility of separated mouse embryos at 2-cell stage, including production of monozygotic twins.
Two-cell embryos were collected from superovulated donor of three types: CF i1 females which were mated with CF # males (#1 embryo), CF #1 females which were mated with C57BL/6 males (F1 embryo) and F1 (CF#1 -C57BL/6) females which were mated with C57BL/6 males (BC embryo). Zona pellucida were removed in Hanks solution containing 0.2-0.5 % pronase. After removal of zona pellucida, the embryos were separated into two halves by pipetting in Ca++ and Mg++ free Hanks solution containing 0.02 EDTA. Separated blastomers were cultured in drops of cultur medium (M16) covered with paraffin oil at 37 °C in an atmosphere of 5% CO2, 95% air for 72 hours. Then a monozygotic pair of half embryos developed into blastocysts were transferred to CF #1 recipient on Day 4 or 5 of pseudpregnancy. The results were as follow: 1) The proportion of single blastomeres developd into blastocysts, for the #1 embryo, Fl embryo and BC embryo were 72.8, 74.2 and 78.5, and the proportion of a monozygotic pair of embryos developed into blastocysts were 61.1, 61.5 and 70.1%, respectively. 2) The cell numbers of half embryos after 72 hours culture, for the #1 embryo, Fl embryo and BC embryo were 51.6± 2.0, 59.0±3.7 and 63.8±2.1, while the cell numbers of normal embryos were 108. 6±5.5, 128.8±3.7 and 136.6 ±4.6, respectively. 3) By transfer of a monozygotic pair of half embryos into recipients, number of pups were obtained 3 (16.7%), including one pair of twins for the Fl embryo and 1 (5.6 %) for the BC embryo. By transfer of a pair of embryo plus eight of normal embryos into recipient, number of pups were obtaind 5 (25.5 %), including 2 pair of twins for the Fl embryo and 1 (3.6%) for the BC embryo. Three sets of monozygot twins were produced.