Microsurgical Blade Research Articles

Pregnancy rates with bisected bovine embryos

A method for producing identical twin calves is described in which 5.5 to 7.5 day bovine embryos were bisected using a fine microsurgical blade. The resulting demi-embryos were transferred surgically or nonsurgically to the uterine horn ipsilateral to the corpus luteum of synchronous recipients (± 2 days), one demi-embryo per recipient. The mean pregnancy rate per demi-embryo was 48% (157/330), and per whole embryo split, it was 95% (157/165). Pregnancy rate was dependent on the stage of embryonic development. The pregnancy rate from bisected early morulae, 16% (7/44), was lower than that from bisected early blastocysts, 60% (58/96) P<0.01. The rate of identical twin production per embryo split reflected these pregnancy rates. Bisected blastocysts resulted in more identical twin sets, 39% (24/62), than early morulae, 9% (2/22) P<0.01. When used with normal embryo transfer procedures, this technique provides a method for routine production of identical twin calves. Furthermore, more calves are produced per two demi-embryos than per whole embryo.

Deep-freezing of cow ‘half’ and ‘quarter’ embryos

This paper describes a simplified method for biopsying goat embryos for sex determination, followed by the transfer of these embryos to recipient does. A 90° angle holding pipet is described, which permits holding the embryo in position and, with minimal effort, open the zona pellucida and excise 5–10 trophoblast cells with the aid of a microsurgical blade. Twenty-four embryos were biopsied, 12 of which were freshly collected (day 7) and 12 of which were cultured in vitro from collected morulae (day 6). In the course of the polymerase chain reaction (PCR) a 447 (male) and 445 bp (female) fragment of the DNA, derived from the trophoblast cells, was amplified. The primers used were specific for the highly conserved mammalian sequences of the ZFX and ZFY genes. Individual biopsied embryos were transferred to 24 synchronized recipient does. The pregnancy rate following transfer was 67% (8/12) in recipients receiving freshly collected blastocysts while only 8% (1/12) in the recipients receiving blastocysts cultured in vitro from morulae (P < 0.01). Four of the kids born were males and four females. The sex assigned to the embryos was correct in 83% of the cases.