• Journal of Embryo Transfer
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• v.16 no.1
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• pp.29-34
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• 2001

To investigate the effect of co-transfer of trophoblastic vesicle (TV) with frozen-thawed in vitro Produced (IVP) bovine embryo on pregnancy rate, IVP blastocysts were transferred to synchronized recipients. Elongated blastocysts were recovered at Day 13 to 15, and dissected more than 4 pieces to removed the embryonic disc. Throphoblastic fragments were cultured for 48 hours to make throphoblastic vesicles (TVs). TVs were cryopreserved in ethylene glycol or vitrification solution and frozen-thawed TVs were co-transferred to recipients with frozen-thawed IVP embryos. 1 The recovery rate of elongated blastocyst on Day 13 to 15 was 22.5% (18/80) and the size of recovered elongated blastocysts was 0.2∼5.0mm. 2. Eighteen elongated blastocysts were dissected into 88 pieces and 61.4% of those pieces were formed to TV (54/88) 3. The viability of frozen-thawed TV in ethylene glycol was higher than in vitrified solution (92.8% vs. 68.8%) 4. The pregnancy rate in co-transfer with frozen-thawed TV and IVP blastocyst was better than transfer only IVP blastocysts (50.0% vs. 23.1%).

• Journal of Embryo Transfer
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• v.25 no.3
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• pp.141-144
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• 2010

In vivo embryo produced from Hanwoo donor cows were collected and transferred to Hanwoo recipients. Cows, at random stages of the estrous cycle, received Progesterone Releasing Intravaginal Device (CIDR-plus, InterAg, New Zealand) together with injection of 1 mg estradiol benzoate and 50 mg progesterone, and gonadotropin treatment began 4 day later. For superovulation, a total of 28 mg FSH was intramuscularly injected twice a day in the way of decreasing doses 4 day (5, 5, 4, 4, 3, 3, 2 and 2 mg). Twenty one Hanwoo donor cows were flushed on day 7 of estrus cycle with same FSH and artificial insemination by the same technicians. Embryos were recovered 7 days after the second insemination by flushing the uterus with Embryo Collection Medium. The results obtained were as follows: The rates of transferable embryos were 50.3%, and 78 fresh embryos at morulae and blastocysts stage were transferred into Hanwoo recipients on day 7 of estrus cycle. The pregnancy rates were first embryo transfer 55.6%, 2nd 62.9% and 3rd 57.9%, respectively. In conclusion, These results suggest that CIDR-based superovulation protocol may be effectively used for production of superior Hanwoo embryos. Also, since it seems the condition of recipient cows greatly affect pregnancy rate, it is very important to evaluate recipient for effective cattle production.

• Journal of Veterinary Clinics
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• v.17 no.1
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• pp.234-237
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• 2000

The dornor, 2 years old, 20kg and mixed breed, was bred naturally on day 1 and day 3 of estrus and eight gastrulae were collected by flushing the uterus of the donor after laparatomy on day 13 after the second mating. The embryos were frozen by programmable freezer and preserved for about 3 months in liquid nitrogen. Another bitch in natural estrus, 2 years old, 30kg, mixed breed, was selected as the recipient and the frozen embryos(8 gastrulae) were thawed and each 4 embryos were transferred into upper partr of left and right uterine horn, respectively, on day 13 after the proper mating day determined by vaginal smear. The ecipient delivered 6 offspring 48 days after embryo transfer. Of 6 puppies, one was still birth and two puppies died one month after birth. Parentage test was performed by DNA analysis using microsatellite sequences for 3 puppiers, the recipient, the donor, the male dog bred with the donor, and the male dog raised near to the recipient. The markers selected for the test were CXX 873(133-157 base pair) and CXX 894(141-165 base pair). Using primers manufactured according to the markers, the blood samples were processed for polymerase chain reaction and the PCR products were treated for electrophoresis. The three puppies showed identical band to that of recipient, consequently, it was concluded that the puppies were offspring of the recipient mated naturally by the male dog, not the offspring by embryo transfer.

• Journal of Embryo Transfer
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• v.27 no.4
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• pp.253-258
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• 2012

This study was carried out to effects of ethylene glycol concentration, sucrose and culture day of in vitro production embryo on slow-down freezing in Hanwoo. 6, 7, 8 and 9 day embryos produced in vitro were frozen using 1.8M EG+0.1M sucrose, 1.8M EG+0.5% BSA and 1.5M EG+0.1M sucrose media. Survivability was confirmed after frozen-thawed 24 and 48h and ICM, TE cell number were counted by Hoechst 33342 and PI staining after frozen-thawed 24h. As a result, 1.8M EG+0.1M sucrose group was most significantly (p<0.05) higher compared with the other treatment groups on survivability, TE and total cell number after frozen-thawed 24h ($94.2{\pm}2.6%$, $94.67{\pm}3.4$ and $129.67{\pm}5.5$). ICM number did not found significant (p<0.05) differences between the three treatment groups. in 6, 7, 8 and 9 day of embryos using three types of freezing media, frozen-thawed, 1.8M EG+0.1M sucrose groups with embryos cultured 8 day was significantly (p<0.05) highest survivability to $98.3{\pm}1.7%$ after frozen-thawed 24h. 1.5M EG+0.1 sucrose group with embryos cultured 9 day was significantly higher survivability than group of embryos cultured 8 day after frozen-thawed 24 and 48h. In conclusion, 1.8M EG+0.1M sucrose media is considered to be effective to cryopreservation of embryos cultured 8 and 9 day.

• Journal of Embryo Transfer
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• v.12 no.1
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• pp.103-110
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• 1997

The present study was carried out to investigate effects of gonadotropin, age of donor, day of estrus cycle gonadotropin injection started and season on embryo production after superovulation in dairy cattles. Embryo collection records were obtained from 177 embryo donor collections from 98 Holstein cows aged from 3 to 9 years during 4 years(1993~1996) at National Livestock Re-search Institute. Superovulation was induced by injections of 3 gonadotropins(FSH-P, FOLLTROPHIN-v* or SUPER-OV*) beginning on days 9 to 14 of the estrus cycle. Em-bryos were collected from donors using a nonsurgical technique on days 6 to 8 after insemi-nation. The results were as follows ;Number of total and freezable embryos per donor cow was affected by gonadotropin(P<0.01). The more number of total and freezable embryos wereobtained by use of FOLLTROPHIN-V (13.2, 7.4) or FSH-P (11.0, 5.7) than SUPER-OV* (5.0, 2.4). Age of donor, the day that gonadotropin was started or season didn't affect total or freezable embryos(P >0.05).

• Journal of Embryo Transfer
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• v.32 no.4
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• pp.275-285
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• 2017

Lysophosphatidic acid (LPA) is an important signaling molecule. Here, the effect and mechanism of LPA on the preimplantation development of porcine embryos during in vitro culture (IVC) was examined. Porcine embryos were cultured in porcine zygote medium (PZM-3) supplemented with $30{\mu}M$ LPA during different days. There was a significantly higher cleavage rate in Day 1-7 and significantly higher total cell number of blastocysts in Day 1-3 and Day 4-7. It was also found that messenger RNA (mRNA) expression level of PCNA, BCL2 and BAX in blastocysts obtained from D1-7 group were significantly higher and BCL2/BAX mRNA ratio in D1-3 group was significantly lower than control group but Day 4-7 and Day 1-7 groups were comparable with control group. Treatment with $20{\mu}M$ PLC inhibitor significantly decreased the embryo cleavage rate and blastocyst formation rate. Moreover, LPA as an activator of PLCs, enhanced the $30{\mu}M$ LPA + $20{\mu}M$ U73122 group embryo cleavage rate which similar with control group. In conclusion, the results suggest that treatment with LPA during IVC improves the porcine early embryo cleavage by activation of PLC signaling pathway and regulate the mRNA expression that contribute to total cell number of blastocysts during blastocyst formation.

• Journal of Embryo Transfer
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• v.4 no.1
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• pp.41-45
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• 1989

Total thirty of flushing were attempted on day 4 to 15 of estrus cycle with S heifers and 9 cows by nonsurgical method. The flushed or recovered rate among flushings was 86.7% (26/30) or 88.5% (23/26), respectively. There was no difference in the recovered rate between heifers (85.7%,6/7) and cows (89.5%, 17119). The embryo was recovered on day 4 to 15 of estrus cycle from the donors in natural heat without any technical difficulties.The I2FG Foley catheter used for pubertal heifers had sometimes plug in it with uterine mucus during flushing of uterine horn. But the problem could be overcomed by pumping the catherter with fluthing solution or by changing the catheter. Three normal embryos were recovered from 3 pubertal (10-11 month old) heifers. The rate of normal and abnormal eggs was 60.9% (14123) and 39.1% (9/23), respectively. The abnormal eggs were on degenerating except one unfertilized egg and were mostly recovered from heifers or cows flushed consecutively during the estrus cycle. The developmental states of normal embryos were l6-cells on day 5, 32-cells on day 6, compacted-morula on day 7, early-to expanded-blastocyst on day 8-to 9, and hatching-to hatched-blastocyst on day 10 to 11 of estrus cycle. The stage of embryos on day 8 to 10 showed varities among donors. On day 8 to 9 of estrus cycle hatching-blas tocyst was recovered from some donors.

• Journal of Embryo Transfer
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• v.5 no.2
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• pp.38-44
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• 1990

This study was carried out to produce superior dairy cattle by embryo transfer. Seven dairy cows were superovulated with divided injection of FSH 4Omg for 5 days started on day 9 to 14 of the estrus cycle and injection of PGF$_2$$\alpha$ 45mg on day 4 of FSH injection. Donor cows were flushed to collect embryos on day 7 or 8 of the estrus cycle. Fresh embryos collected were transferred to synchronized dairy recipients or frozen using glycerol 3 step method to he equilibrated. And 35 embryos which were frozen using glycerol 6 step method were imported from U.S.A. After glycerol dilution of frozen embryos was done by reverse density during freezing. frozen-thawed embryos were transferred to synchronized dairy or beef recipients. The results obtained were as follows; 1. Total of 24 embryos were collected from 7 donor cows flushed and transferable embryos were 18 (75.0%). 2. Among 24 embryos. morula, early blastocyst, blastocyst, expanded blastocyst and unfertilized ova were 3 (12.5%), 1 (4.2%), 10 (41.6%), 4 (16.7%) and 6 (25.0%), respectively. 3. Heat inducing rate after 1st and 2nd injections of PGF$_2$$\alpha$ in Holstein and beef cattle was 83.3% and 71.4% and 62.5% and 69.2%, respectively. 4. Among 56 recipients, 23 head were pregnant (41.1%). The pregnancy rate of fresh embryos was 50.0% (1/2 heads) and the pregnancy rate of frozen embryos which were frozen using glycerol 3 step and using glycerol 6 step imported from U.S.A. was 52.6%(l0/19 heads) and 34.3%(12/35 heads), respectively. 5. The pregnancy rate of blastocyst (60.0%) was higher than that of morula (39.0%), early blastocyst (25.0%) and expanded blastocyst (0%). 6. The pregnancy rate of grade I embryos (52.2%) was higher than that of grade 2 (34.6%) and grade 3 (28.6%). 7. The pregnancy rate according to synchrony of recipient with donor was higher in simultaneous recipient (55.0%) and +l2hrs' (53.8%) than -24hrs' (23.5%), -l2hrs' (20.0%) and +24hrs' (0%).

• Journal of Embryo Transfer
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• v.6 no.2
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• pp.37-46
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• 1991

A field trial was performed to evaluate the effects of hormone treatment on estrus induction, ovulation, embryo transfer and reproductive performance in post-weaning sows. This trial involved 61 mixed breed sows of varying parity on a commercial pig farm. Sows were allocated to one of five trials: control group involved 25 sows that were treated with a single intramuscular injection of 5 ml physiological saline, 6 sows received 1,500 IU PMSG on the day of weanning and 500 IU HCG at the onset of estrus in trial I, 7 sows received 750 IU PMSG on the day of weanning and 500 IU HCG at the onset of estrus in trial II, 5 sows were treated with the same as trial II on day 28 after weanning in trial III. and 18 sows were treated with 10 mg PGF$_2$$\alpha$ plus 2 mg estradiol benzoate on day 31 after weanning in trial IV. Ovarian responses were checked by laparotomy and ova were recovered by oviducal flushing between 40 and l00hrs after mating. Fertilized ova were transferred into the oviduts of recipient sows synchroni- zed. The results obtained were summarized as follows: 1. Percentages of sows detected in standing estrus following treatment were 86~100% among trial groups. The interval from treatment to standing estrus(6l.7$\pm$0.5lhrs) in lOmg PGF$_2$$\alpha$ and 2mg estradial henzoate treated group was significantly earlier than in other trial groups(P<0.05). 2. Average number of ovulations was 11.5~37.8 among trial groups. The ovulation rate in 1,500 IU PMSG and 500 IU RCG treated group (37.8$\pm$ 19.87) was significantly different from other trial groups(P<0.05). 3. Ova were recovered by oviducal flushing between 40~ l00hrs after mating and recovery rates of ova wore 91.4% between 40~59hrs. 4. Fertilized ova were transferred into the oviducts of 8 recipient sows synchronized with 7 to 17 ova per animal. Three of the recipients were pregnant and delivered 25 piglets. 5. Four of the donor sows in those embryo collection was not successful were pregnant following oviducal flushing and delivered 23 piglets. 6. Recurrence of estrus and farrowing performance of experimental sows were observed following the experiment was no difference among trial groups, respectively.

• Journal of Embryo Transfer
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• v.20 no.2
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• pp.89-95
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• 2005

In this study, we investigated the effects of number, grade, stage and day of embryos on pregnancy and abortion rates of Holstein heifer recipients after transfer of in-vitro produced Korean Native Cattle (Hanwoo) embryos. When one, two or three blastocysts were transferred to recipients, pregnancy (32.0, 44.0 and $32.0\%$) and abortion (12.5, 13.6 and $6.3\%$) rates did not differ among treatments. The pregnancy and abortion rates of different grades of blastocysts were $40.8\~44.2\%\;and\;9.4\~16.3\%$, respectively. However, they were not significantly different. According to the stage of blastocyst, the pregnancy rates were similar between $39.1\%\;and\;45.3\%$. However, the abortion rate in hatched blastocyst stage $(20.8\%)$ was significantly higher than that in late blastocyst stage $(9.0\%)$. The pregnancy rates in day $7\cdot7$ and day $7\cdot8$ blastocyst (47.1 and $53.9\%$, respectively) was significantly higher than that in the day $9\cdot9\;and\;8\cdot9$ blastocyst (29.8 and $27.7\%$, respectively). The abortion rate was the highest in the day $9\cdot9$ blastocyst $(30.8\%)$ among treatments, and it was significantly different compared to that in the day $7\cdot7$ blastocyst $(10.1\%)$. Therefore, this study suggested that the pregnancy rate was affected by only day of embryos, while the abortion rate was affected by stage and day of embryos.