Production of monozygotic mouse twins from microsurgically bisected morulae

Abstract

Mouse monozygotic twins were produced by bisection of the compacted morulae and transfer of the pairs of half-embryos after culture in vitro. The compacted morulae (about 16 cells) were microsurgically bisected, using a fine glass needle attached to a micromanipulator, without any supporting micro-instruments, after pretreatment for zona-softening and decompaction. About 80% of the morulae were bisected without visible cell damage. After 20 h in culture, the half-embryos were classified morphologically as eu-blastocysts, pseudo-blastocysts, or trophectodermal vesicles or non-integrated forms. After culture of 131 pairs of bisected morulae, 75 (57.3%) pairs of eu-blastocysts, 20 (15.3%) pairs comprising a eu-blastocyst and pseudo-blastocyst, and 9 (6.9%) pairs of pseudo-blastocysts, were obtained. The pseudo-blastocysts were considered to be derived from half-morulae in which some blastomeres were destroyed or dissociated as a result of micromanipulation. From 30 pairs of eu-blastocysts transferred to 21 recipients, 5 twin fetuses on Day 17 (18 pairs/9 recipients) and 3 twin male young (12 pairs/12 recipients) were obtained. Survival rate of the twin-embryo pairs was 27.8% at autopsy and 25.0% at term. None of the 20 pairs of pseudo-blastocysts transferred to 10 recipients gave rise to normal conceptuses.