IN-VITRO EMBRYO PRODUCTION AFTER MICROINJECTION AND OVARIAN DYNAMICS FOLLOWING TRANSVAGINAL, FOLLICULAR OOCYTE ASPIRATION

Ultrasound-guided transvaginal follicular aspiration of oocytes from l ive cows combined with IVM, IVF and in vitro culture (IVC) is a proced ure for producing preimplantation-stage bovine embryos and a source of oocytes for pronuclear microinjection of DNA for producing transgenic cattle. This experiment was designed to compare in vitro embryo devel opment rates between oocytes derived from transvaginal follicular aspi ration and those obtained from cows at slaughter. Nine cows were subje ct to a twice-weekly aspiration. Oocytes were aspirated with a 5 MHz u ltrasound transducer packaged in a vaginal probe equipped with a dorsa l-mounted needle guide (16-ga). All visible follicles (>2 mm) were pun ctured with a 17-ga, 55-cm needle at each aspiration session and the c ontents removed under vacuum suction. Oocytes underwent IVM/IVF/IVC. M icroinjection of DNA was performed during the pronuclear stage of deve lopment, and the zygotes were co-cultured on Buffalo Rat Liver (BRL) c ells in modified M199 at 39 degrees C in 5% CO2 and air. After 7 d in culture, embryos were removed and scored for development. A Chi-square analysis was used to compare transvaginal follicular-derived oocytes (microinjected and not) and slaughterhouse-derived, matured in transit oocytes (SHDMT; microinjected and not). Nonmicroinjected embryos resu lting from IVF of transvaginal aspiration-derived oocytes developed to blastocysts at a higher rate than SHDMT oocytes (40.0 vs 30.8%; P<0.0 5). There was no difference in development rates between the microinje cted groups (aspiration = 15.9% vs SHDMT = 12.8%). Higher proportions of the embryos generated from the aspirated oocytes were of excellent or good quality following culture (P<0.05). In the present experiments the effects of microinjection may overshadow some effects of ova sour ce, but transvaginal follicular aspiration may provide a more consiste nt, synchronous population of oocytes than those derived from commerci al slaughter house sources for use with in vitro systems.