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bioRxiv · 10.1101/2022.02.18.481106

SUCCESSFUL IN VITRO EMBRYO PRODUCTION WITH OOCYTES ASPIRATED FROM LIVE WHITE-TAILED DEER (Odocoileus virginianus texanus) DONNORS UNDER CAPTIVITY IN NORTHEAST MEXICO

MarabotoM., E. E.·Trejo M., F. J.·Del Angel R., H.·Rosales H., J.·Bautista M., Y.·Torres P., L. I.·Gonzalez-Reyna, A.
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Abstract

Assisted reproductive technologies (ARTs), such as artificial insemination, semen sorting and freezing, embryo production in vivo and in vitro, are all methods in which animal reproduction management of several species has become considerably more efficient. These ARTs have been applied both in domestic and wild ruminants. In this study, in vitro embryo production was attempted with oocytes collected surgically during the mating season, from live white-tailed deer (WTD) hinds, maintained under captivity, in Northeast Mexico. The study was conducted in two WTD farms nearby Cd. Victoria, Tamps., Mexico, and another WTD farm located in Guerrero Coahuila, Mexico. The laboratory work was carried at the Centro de Desarrollo de la Capacidad Productiva y Mejoramiento Genetico de la Ganaderia, property of the Union Ganadera Regional de Tamaulipas (Centro-UGRT). The deer hinds are kept in captivity year-around, with feeding and health management provided accordingly to their requirements (50-60 kg females). Fresh green forage and clean fresh water is supplied daily. Oocytes were collected on each farm, while the hinds were maintained under general anesthesia and by means of a mid-ventral laparotomy, ovaries were exposed and follicles greater than 1-2 mm were aspirated with a 20 G needle, connected by a two-way plastic and latex hose to a vacum machine (WTA, Brazil); the oocytes were collected into a 50 ml. centrifuge tube containing wash media with heparine (Vitrogen, Brazil). Each oocyte collection lasted approximately 20 minutes, after which and while still on the farm, the oocytes were filtered with 50 {micro} mesh filters and rinsed several times with wash media, then placed on search Petri plates. Oocytes were then counted and classified (total, viable and non-viable oocytes) and placed into cryovials with maturation media (3 ml. MIV, Vitrogen, Brazil) and placed into a portable incubator with 5 % CO2 gas mix (LabiMixWTA, Brazil), after all hinds were done, oocytes were transported to the in vitro fertilization laboratory (IVF Lab) at the Centro-UGRT. Once at the IVF Lab, the oocytes were placed on a larger incubator (Eve, WTA, Brazil) and kept there for 18-20 hours, after which, the MIV media was changed for fertilization media (FIV, Vitrogen, Brazil) and fertilization was initiated by adding 10,000-12,000 live sperm per cryovial, and incubated for an additional period of 24 hours; after which, FIV media was replaced by the same media and at this point, cleavage rate was estimated by counting the oocytes that initiated cell division. At 72 hours after cell division started, fertilization rate was estimated; and 7 days after, the blastocysts were counted and classified. The whole process of oocyte maturation and embryo production in vitro, was conducted based on a beef cattle embryo production system and adapted to a deer embryo production system using media for small ruminants (Vitrogen, Brazil). Data collected per hind included total number of oocytes, viable and non-viable oocytes, cleavage rate (ratio of viable oocytes that initiated cell division over viable oocytes), fertilization rate (ratio of embryos that initiated cleavage over those that continued development to the blastocyst stage) and blastocyst rate (embryos reaching the blastocyst stage over cleaved embryos); averages were calculted for each parameter. The main results from this study on a per hind basis for total viable and non-viable oocytes were 9.8, 6.3 and 4.5, respectively; cleavage and blastocyst rates were 39.5 and 36.8, respectively and 2.3 blastocysts. In conclusion, oocyte collection from live WTD hinds and in vitro embryo production was succesfully done under farming conditions in Northeast Mexico.

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MarabotoM., E. E., Trejo M., F. J., Del Angel R., H., Rosales H., J., Bautista M., Y., Torres P., L. I., Gonzalez-Reyna, A.. 2022-02-25. SUCCESSFUL IN VITRO EMBRYO PRODUCTION WITH OOCYTES ASPIRATED FROM LIVE WHITE-TAILED DEER (Odocoileus virginianus texanus) DONNORS UNDER CAPTIVITY IN NORTHEAST MEXICO. https://doi.org/10.1101/2022.02.18.481106

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