RESUMO

Artificial insemination (AI) was the first important biotechnology applied to improve the genetics of farm animals. It allows the rapid and massive diffusion of desirable characteristics of males with high productive potential. We describe the different types of estrus induction and synchronization techniques and the use of the AI with fresh, chilled or frozen semen. Through the adequacy of the protocols of estrus synchronization and AI to the different production systems, the efficient use of reproductive techniques is possible, reaching acceptable pregnancy rates. Summary of reproductive results obtained using cervical and laparoscopic AI are presented.

RESUMO

The aim was to evaluate pregnancy success after transfer of embryos vitrified in micropipette tips in Merino sheep under extensive conditions. A second objective was to evaluate the influence of embryo stage in such pregnancy rate. One hundred and twenty-seven embryos were rewarmed and transferred into recipient ewes. On rewarming, the embryos were placed into three-step cryoprotectant dilutions. Finally, prior to transfer to recipient females, embryos were maintained in Basic Medium for 5 min at 25ºC and were re-evaluated by morphological criteria; all degenerated embryos were eliminated. Recipient ewes (n = 150) were treated for estrus with sponges placed for 14 days and 300 IU of eCG. At embryo transfer, three experimental groups were defined: morulae transferred on Day 7, blastocysts transferred on Day 7 and blastocysts transferred on Day 8 after sponge removal. In all groups, semi-laparoscopic transfer of one rewarmed embryo per recipient was performed. Pregnancy was diagnosed by ultrasonography on day 28 after embryo transfer. The embryo selection rate after rewarming was higher for blastocysts (89.3% - 67/75) compared to morulae (65.9% - 60/91) (P < 0.05). Pregnancy diagnosis showed a 38.3% (23/60) of success after morula transfer on Day 7 post progestagen removal. The day of transfer showed a significant influence on pregnancy rate after blastocyst transfer (Day 8, 55.9% - 19/34 vs Day 7, 21.2% - 7/33) (P < 0.05). Blastocysts transfer on Day 8 showed the highest global efficiency (pregnancies/total embryos after rewarming) (47.5% - 19/40) (P < 0.05). In conclusion, reproductive efficiency obtained by vitrified embryo transfer allows its recommendation for embryo transfer programs under extensive conditions. The importance of considering the synchrony between the embryo age and the recipient uterus stage is emphasized.

RESUMO

The aim was to evaluate pregnancy success after transfer of embryos vitrified in micropipette tips in Merino sheep under extensive conditions. A second objective was to evaluate the influence of embryo stage in such pregnancy rate. One hundred and twenty-seven embryos were rewarmed and transferred into recipient ewes. On rewarming, the embryos were placed into three-step cryoprotectant dilutions. Finally, prior to transfer to recipient females, embryos were maintained in Basic Medium for 5 min at 25ºC and were reevaluated by morphological criteria; all degenerated embryos were eliminated. Recipient ewes (n = 150) were treated for estrus with sponges placed for 14 days and 300 IU of eCG. At embryo transfer, three experimental groups were defined: morulae transferred on Day 7, blastocysts transferred on Day 7 and blastocysts transferred on Day 8 after sponge removal. In all groups, semi-laparoscopic transfer of one rewarmed embryo per recipient was performed. Pregnancy was diagnosed by ultrasonography on day 28 after embryo transfer. The embryo selection rate after rewarming was higher for blastocysts (89.3% - 67/75) compared to morulae (65.9% - 60/91) (P < 0.05). Pregnancy diagnosis showed a 38.3% (23/60) of success after morula transfer on Day 7 post progestagen removal. The day of transfer showed a significant influence on pregnancy rate after blastocyst transfer (Day 8, 55.9% - 19/34 vs Day 7, 21.2% - 7/33) (P < 0.05). Blastocysts transfer on Day 8 showed the highest global efficiency (pregnancies/total embryos after rewarming) (47.5% - 19/40) (P < 0.05). In conclusion, reproductive efficiency obtained by vitrified embryo transfer allows its recommendation for embryo transfer programs under extensive conditions. The importance of considering the synchrony between the embryo age and the recipient uterus stage is emphasized.

Assuntos

RESUMO

Artificial insemination (AI) was the first important biotechnology applied to improve the genetics of farm animals. It allows the rapid and massive diffusion of desirable characteristics of males with high productive potential. We describe the different types of estrus induction and synchronization techniques and the use of the AI with fresh, chilled or frozen semen. Through the adequacy of the protocols of estrus synchronization and AI to the different production systems, the efficient use of reproductive techniques is possible, reaching acceptable pregnancy rates. Summary of reproductive results obtained using cervical and laparoscopic AI are presented.

Assuntos

RESUMO

The aim was to evaluate pregnancy success after transfer of embryos vitrified in micropipette tips in Merino sheep under extensive conditions. A second objective was to evaluate the influence of embryo stage in such pregnancy rate. One hundred and twenty-seven embryos were rewarmed and transferred into recipient ewes. On rewarming, the embryos were placed into three-step cryoprotectant dilutions. Finally, prior to transfer to recipient females, embryos were maintained in Basic Medium for 5 min at 25ºC and were reevaluated by morphological criteria; all degenerated embryos were eliminated. Recipient ewes (n = 150) were treated for estrus with sponges placed for 14 days and 300 IU of eCG. At embryo transfer, three experimental groups were defined: morulae transferred on Day 7, blastocysts transferred on Day 7 and blastocysts transferred on Day 8 after sponge removal. In all groups, semi-laparoscopic transfer of one rewarmed embryo per recipient was performed. Pregnancy was diagnosed by ultrasonography on day 28 after embryo transfer. The embryo selection rate after rewarming was higher for blastocysts (89.3% - 67/75) compared to morulae (65.9% - 60/91) (P < 0.05). Pregnancy diagnosis showed a 38.3% (23/60) of success after morula transfer on Day 7 post progestagen removal. The day of transfer showed a significant influence on pregnancy rate after blastocyst transfer (Day 8, 55.9% - 19/34 vs Day 7, 21.2% - 7/33) (P < 0.05). Blastocysts transfer on Day 8 showed the highest global efficiency (pregnancies/total embryos after rewarming) (47.5% - 19/40) (P < 0.05). In conclusion, reproductive efficiency obtained by vitrified embryo transfer allows its recommendation for embryo transfer programs under extensive conditions. The importance of considering the synchrony between the embryo age and the recipient uterus stage is emphasized.(AU)

Assuntos

RESUMO

Artificial insemination (AI) was the first important biotechnology applied to improve the genetics of farm animals. It allows the rapid and massive diffusion of desirable characteristics of males with high productive potential. We describe the different types of estrus induction and synchronization techniques and the use of the AI with fresh, chilled or frozen semen. Through the adequacy of the protocols of estrus synchronization and AI to the different production systems, the efficient use of reproductive techniques is possible, reaching acceptable pregnancy rates. Summary of reproductive results obtained using cervical and laparoscopic AI are presented.(AU)

Assuntos

RESUMO

Transgenesis constitutes an important tool for pharmacological protein production and livestock improvement. We evaluated the potential of laparoscopic insemination (LI), in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) to produce egfp-expressing ovine embryos, using spermatozoa previously exposed to pCX-EGFP plasmid in two different sperm/DNA incubation treatments: "Long Incubation" (2 h at 17 C) and "Short Incubation" (5 min at 5 C). For LI, Merino sheep were superovulated and inseminated with treated fresh semen from Merino rams. The embryos were recovered by flushing the uterine horns. For IVF and ICSI, slaughterhouse oocytes were fertilized with DNA-treated frozen/thawed sperm. All recovered embryos were exposed to blue light (488 nm) to determine green fluorescent morulae and blastocysts rates. High cleavage and morulae/blastocysts rates accompanied the LI and IVF procedures, but no egfp-expressing embryos resulted. In contrast, regardless of the sperm/plasmid incubation treatment, egfp-expressing morulae and blastocysts were always obtained by ICSI, and the highest transgenesis rate (91.6%) was achieved with Short Incubation. In addition, following the incubation of labeled plasmid DNA, after Long or Short exposure treatments, with fresh or frozen/thawed spermatozoa, only non-motile fresh spermatozoa could maintain an attached plasmid after washing procedures. No amplification product could be detected following PCR treatment of LI embryos whose zonae pellucidae (ZP) had been removed. In order to establish conditions for transgenic ICSI in the ovine, we compared three different activation treatments, and over 60% of the obtained blastocysts expressed the transgene. For ICSI embryos, FISH analysis found possible signals compatible with integration events. In conclusion, our results show that in the ovine, under the conditions studied, ICSI is the only method capable of producing exogenous gene-expressing embryos using spermatozoa as vectors.

Assuntos