• Journal of Embryo Transfer
• /
• v.26 no.4
• /
• pp.271-276
• /
• 2011

This study was focused on improvement of milk production in Mongolian dairy industry by artificial insemination (AI) technology, supported by ODA of KOICA in Republic of Korea. This program was started in January 2009 and it is in $3^{rd}$ years. This manuscript summarized the data especially on estrus synchronization and pregnancy establishment in dairy cows (Holstein) this year. A total of 81 dairy cows from 4 private farms (38 from Undarmal milk and that of 30, 8 and 5 dairy cows from Onjin (Enkhbayer), Jargalant, and BRM School farms respectively) were synchronized with 5 ml Lutalyse (i.m.) in the dump of dairy cows and then estrus was detected 2 to 3 days after $PGF_{2{\alpha}}$ injection. The synchronized dairy cows were inseminated with 0.5 ml dairy frozen semen by conventional artificial insemination (AI) techniques. Pregnancy was diagnosed about 60 days after AI by palpation method. About 96.3% (78/81) of synchronized cows were responded to single $PGF_{2{\alpha}}$ injection. Total 75 over 78 dairy cows (90.1%) inseminated were diagnosed as pregnant. The estrus induction and pregnancy rates were very effective using Lutalyse injection and conventional AI techniques in Mongolian dairy cow. The present results indicated that AI after estrus induction in Mongolian dairy cows could be applied to dairy breeding technology for improving breeding efficiency and milk production of the country.

• Journal of Embryo Transfer
• /
• v.17 no.1
• /
• pp.1-5
• /
• 2002

The purpose of this study was carried out to determine the possibility of continued embryo collection by repeatable superovulation from a Hanwoo elite cow, who was already estimated to an elite cow by DNA analysis. The donor cow was superovulated with Folltropin-V and PG $F_2$$\alpha$ combination method for 5 times, 2 months interval during 10 months. The estrus cows were inseminated with frozen semen 3 times 12 hrs interval. Embryo collection from superovulated cows was flushed non-surgical method at 7 to 8 days after artificial insemination. The superovulated cow was collected from 1 to 5$^{th}$ times as total (14, 4, 10, 6, 9 embryos; total 30 embryos) and transferable embryos (7, 4, 10, 3, 6; Mean$\pm$S.E : 6.0$\pm$2.7 embryos). Seventeen recipients were transferred 24 embryos and then detected to pregnant 10 recipients after embryo transfer (58.8%; 10/17 recipients) The results indicated that the possibility of repeatable superovulation from elite donor cow for embryo transfer could be confirmed in this study, although the replicated number was not enough.

• Journal of Animal Science and Technology
• /
• v.44 no.3
• /
• pp.279-288
• /
• 2002

This study was performed to discriminate defective loci by detection of congenital genetic disorder, to offer basic data for selection and improvement of Korean dairy cattle using frozen semen of Holstein bulls(16 proven and 93 candidate). The results obtained were as follows ; By the detection of DUMP(deficiency of uridine monophophate synthase) for 109 Holstein bulls(16 proven and 93 candidate), DUMP carrier was not found in whole animals. Also, it was possible to early detection of DUMP carrier by using PCR-RFLP(AvaⅠ). As the results of detection for BLAD(bovine leukocyte adhesion deficiency), BLAD carrier was not found in 16 proven bulls. But 5 candidtae bulls are discriminated to BLAD carrier, and it could be predicted to transmitted pathway of inherited loci by pedigree identification. Also, when digesting PCR products using restriction enzyme, results from TaqⅠ restriction enzyme were more efficient than that of HaeⅢ. After detection test of citrullinaemia, it was concluded that proven and candidate bulls were not. However, wide range of research and citrullinaemia genotyping should be performed. As a result of this study, the wide and various research should be performed in genetic disease of animal. And in the selection and breeding of animal, the breeding scheme by completely and continuously management of pedigree should be established.

• Korean Journal of Agricultural Science
• /
• v.14 no.2
• /
• pp.413-421
• /
• 1987

The objective of this study was to evaluate the possibility of sex preselection by gradients methods using bovine serum albumin in rabbits. Artificial insemination with separated sperm was performed, after highly motile sperm were separated by different methods using 6%, 10% and 20% bovine serum albumin. Various characteristics of separated sperm, and the conception rate and secondary sex ratio at artificial insemination with sperm separated by different methods were compared. The results obtained were as follows. 1. The conception rate of sperm separated by bovine serum albumin gradients was higher than th at of control sperm. But secondary sex ratio was not altered by this methods. 2. The sperm separated by bovine serum albumin gradients showed significantly high value in motility, percent of normal sperm and progressive motility as compared with control sperm and revealed the highest sperm recovery when separated with 6% bovine serum albumin. 3. The sperm motility, percent of normal sperm and progressive motility of the highly motile sperm frozen after being separated from raw semen with bovine serum albumin, showed significantly high value than those of control sperm.

• Journal of Veterinary Clinics
• /
• v.30 no.5
• /
• pp.384-386
• /
• 2013

A two-year-old, female English bulldog was referred for breeding by artificial insemination with frozen semen of male English bulldog, a litter of female bulldog's grandfather. Intrauterine artificial insemination was done two days after the ovulation day. Sperm was evaluated after thawing by computer assisted sperm analyzer, and its motility was 89.8% with normal shape. Pregnancy bearing eight fetuses was diagnosed by ultrasonography and radiography. Cesarean section was performed sixty days after the artificial insemination. Eight pups were delivered with safe, but the entire pup had abnormalities including severe bow-legged malformations, cleft lip, cleft palate, and enlarged cranial part.

• Korean Journal of Animal Reproduction
• /
• v.24 no.4
• /
• pp.439-449
• /
• 2000

This study was to investigate the expression of transgene after co-injection of spermatozoon and EGFP gene into mature oocytes in cattle. From frozen semen, spermatozoa were treated by DTT with 0.03% Tween-20, freezing and thawing or 0.02% Triton X-100 to disrupt their plasma membranes. The sperm injected oocytes were co-cultured with bovine oviduct epithelial cells in CRlaa, and expression of EGFP in embryos were observbed under epifluorescent microscope. Two pronuclei were formed in oocytes injected with sperm treated by DTT (44.6%), DTT-Tween-20 (48.4%), DTT-freezing (44.4%) and DTT-Triton X-100 (42.9%). Cleavage and blastocyst formation rates of bovine oocytes which injected with sperm treated by DTT, DTT-Tween-20, DTT-freezing, and DTT-Triton X-100 were 49.1, 58.5, 43.9, and 48.4% and 10.2, 13.0, 6.8, and 6.5%, respectively. Although the most of embryos were showing mosaicism, embryos expressing EGFP gene were observed in all treated groups. Therefore, these results indicate that membrane disrupted sperm could interact with exogenous DNA, and that this procedure may be useful to introduce foreign gene into bovine oocytes.

• Journal of Embryo Transfer
• /
• v.16 no.2
• /
• pp.99-106
• /
• 2001

This study was undertaken to compare the efficiency of recovery rate and development rate of follicular oocytes collected either by aspiration or by slicing method. The follicular oocytes collected by the two methods matured in TCM199 supplemented with 10% steer serum at 39$^{\circ}C$ in a humidified atmosphere of 5% $CO_2$in air. After 22 h of culture, the oocytes were inseminated with frozen-thawed semen (2$\times$10$^{6}$ sperm/ml of final concentration) prepared with Percoll-density gradient in IVF-TALP medium for 16 h. Later, sets of 15 presumptive zygotes were transferred into 50 $\mu$L, droplets of CR1aa medium. On day 4 of the culture, embryos were transferred to TCM199 until day 9. The percentages of nuclear maturation to pre-metaphase II in the oocytes collected by aspiration are significantly (P<0.05) higher than that by slicing (83% vs. 62%, respectively). The mean number of oocytes recovered by slicing per ovary is significantly (P<0.05) higher than that by aspiration (15.1 vs. 6.7, respectively). Although the rates of cleavage and development to blastocyst of oocytes collected b)\\\\`aspiration are significantly (P<0.05) higher than that by slicing, the number of transferable embryos obtained by slicing method is significantly (P<0.05) higher than that by aspiration. From the results. we may conclude that slicing method is better than aspiration method for obtaining large number of transferable embryos per ovary.

• Journal of Embryo Transfer
• /
• v.30 no.3
• /
• pp.121-128
• /
• 2015

The objective of this study was to evaluate the efficiency of sperm cryosurvival in boar sperm separated by Percoll containing antioxidant enzymes. The boar semen was collected into a pre-warmed ($37^{\circ}C$) thermos bottle by gloved-hand method and was separated by 65% Percoll with superoxide dismutase (SOD), catalase (CAT) and glutathione (GSH) before freezing. The frozen sperm was thawed at $38.5^{\circ}C$ for 45 sec in water-bath for sperm characteristic analysis. The sperm were estimated with SYBR14/PI double staining for viability, FITC-PNA/PI double staining for acrosome reaction, Rhodamine123/PI double staining for mitochondrial integrity and were analyzed using flow cytometry. In results, sperm viability, acrosome reaction and mitochondrial integrity were improved in separated sperm groups compared with unseparated sperm by Percoll (UP) group. Especially, viability was significantly higher in sperm separated by Percoll containing 400 IU CAT group compared with other groups (P<0.05). And acrosome reaction was decreased in sperm separated by Percoll with 300 IU SOD, 400 IU CAT and 0.5 mM GSH groups compared with other groups, however, there were no significantly difference mitochondrial integrity among sperm separated by Percoll with antioxidant enzymes. In conclusion, we suggest that use of Percoll containing antioxidant enzymes for sperm separation will be beneficial for sperm cryopreservation in pigs.

• Korean Journal of Veterinary Research
• /
• v.42 no.1
• /
• pp.35-43
• /
• 2002

Cryopreservation is commonly used for an efficient utilization of semen, oocytes and embryos but has disadvantage in the survival, development of the post-thawed eggs. The high risk in the survival, development of eggs after thawing is thought to be caused by inappropriate internal regulation of $Ca^{2+}$ and/or formation of intracellular ice crystals. In this experiment, we tested whether the $Ca^{2+}$ current (iCa), a decisive factor to $Ca^{2+}$ entry, was altered in post-thawed oocytes by using whole cell voltage clamp technique. The quality and survival rates of the oocytes derived from both fresh and frozen groups were examined by morphology and FDA-test. Vitrified oocytes (VOs) were incubated for 4 hr after thawing and then donated to this experiment. Ethyleneglycol-ficoll-galactose (EFG) was used as a cryoprotectant for vitrification. The membrane potential was held at -80 mV and step depolarizations of 250 ms were applied from -50 mV to 50 mV in 10 mV increments. The survival rates showed a higher in VOs vitrified with EFG containing $Ca^{2+}$ than in VOs vitrified with EFG under the $Ca^{2+}$-free condition (82.0% vs 14%). In group with/without $Ca^{2+}$, the survival rates were significantly (P<0.01) difference. In the fresh metaphase II oocytes (FOs), current-voltage (I-V) relationship showed that iCa began to activate at -40 mV and reached its maximum at -10 mV. With same voltage pulses, inward currents were elicited in VOs. I-V relationships observed in VOs were similar to those in FOs. Time constants of activation and inactivation of the inward current shown in VOs were not different to those in FOs. This accordance in I-V relations and time constants in FOs with those in VOs indicates that the inward currents in FOs are unaltered by vitrification and thawing. Therefore, vitrification with EFG does not play as a factor to deteriorate $Ca^{2+}$ entry across the membrane of the oocytes.

• Korean Journal of Veterinary Research
• /
• v.36 no.4
• /
• pp.919-927
• /
• 1996

In the last few years, methods for in vitro culture of early embryo stages from oocytes matured and fertilized in vitro using suitable cell culture systems have been established. But the factors affecting pregnancy rates following transfer of bovine embryos produced in vitro were not evaluated enough. So this study was performed to investigate the effects of quality and stage of embryos, parity and Corpus Luteum quality of recipients on pregnancy rates following non-surgical transfer of bovine embryos produced in vitro. Oocytes aspirated from small antral follicles of ovaries obtained at a local slaughter house were matured, fertilized with frozen-thawed semen and co-cultured for 6-7 days by utilizing co-culture system with bovine oviduct epithelial cell in vitro. After co-culture, embryos were transfered to recipients on day 7 (estrus=day 0). Recipients were monitored by ultrasonic scanning method or observation for estrus and rectal palpation after 50 days from transfer. The results of this study are follows. 1. Of the 70 recipients, 70%(49 of 70) had not showed estrus sign between day 0 and day 50, but 22.9%(16 of 70) was diagnosed not pregnant. Therefore the overall pregnancy rate of this study was 47.1%(33 of 70). 2. The pregnancy rate of recipients transfered with excellent(66.7%) and good(54.5%) embryos were higher than that of recipients transfered with fair embryos(15.8%) (p<0.05). 3. The pregnancy rate of recipients transfered with morula, compacted morula, blastocyst and expanded blastocysts were 46.2, 55.0, 62.5 and 50.0%, respectively. 4. The pregnancy rates of recipients transfered to heifer and cow were 54.5 and 55.2%, respectively. 5. The pregnancy rates of recipients with CL score I, II(66.7, 63.6%) were higher than those of recipients with CL score III (10%), (p<0.05). Success of transfer of embryos produced in vitro depends on many variables. The important factors identified in this study were the quality of embryos and the CL score of recipient animals after non-surgical transfer of embryos matured, fertilized and cultured in vitro.