• Asian-Australasian Journal of Animal Sciences
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• v.16 no.7
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• pp.1071-1086
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• 2003

Reproductive biotechnologies continue to be developed for genetic improvement of both river and swamp buffalo. Although artificial insemination using frozen semen emerged some decades back, there are still considerable limitations. The major problem appears to be the lack of efficient methods for estrus detection and timely insemination. Controlled breeding experiments in the buffalo had been limited and similar to those applied in cattle. Studies on multiple ovulation and embryo transfer are essentially a replica of those in cattle, however with inherent problems such as lower number of primordial follicles on the buffalo ovary, poor fertility and seasonality of reproduction, lower population of antral follicles at all stages of the estrous cycle, poor endocrine status and a high incidence of deep atresia in ovarian follicles, the response in terms of transferable embryo recovery has remained low with 0.51 to 3.0 per donor and pregnancy rates between 15 to 30%. In vitro production of buffalo embryos is a valid alternative to recovery of embryos by superovulation. This aspect received considerable attention during the past decade, however the proportion of embryos that develops to the blastocyst stage is still around 25-30% and hence the in vitro culture procedures need substantial improvement. Embryo cryopreservation procedures for direct transfer post thaw need to be developed for bubaline embryos. Nuclear transfer and embryo cloning is a technique that has received attention in various species during recent years and can be of immense value in buffaloes as they have a low rate of embryo recoveries by both in vitro and in vivo procedures. Gender pre-selection, genome analysis, gene mapping and gene transfer are a few of the techniques that have been studied to a limited extent during recent years and are likely to be included in future studies on buffaloes. Very recently, reproductive biotechnologies have been applied to feral buffaloes as well, but the results obtained so far are modest. When fully exploited they can play an important role in the preservation of endangered species.

• Asian-Australasian Journal of Animal Sciences
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• v.13 no.5
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• pp.595-599
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• 2000

Fine wool sheep (n=18) maintained in a tropical environment were allocated to three treatment groups. Estrus was induced with two injections of $PGF_{2{\alpha}}$ (10 mg. im) at 10 days interval. Superovulation treatment started 2 days prior to the second injection of $PGF_{2{\alpha}}$. Each ewe was treated with a total dose of 25 units FSH (Super-OV) i.m. every 12 hover 3 days; Group 2 were also injected i.m. with 200 IU PMSG at the first injection of FSH; Group 3 was treated as in Group 2 and also with GnRH ($4{\mu}g$ Buserelin) at the onset of estrus. The ewes in estrus were mated with a fertile ram. Ovarian examination and recovery of embryo and ova were performed at laparoscopy and laparotomy on day 3 or 4 after mating. Data for onset of estrus, duration of estrus, number of corpora lutea (CL), number of unnovulated large follicle (LF), embryo recovery rate, embryo quality and fertilization recorded for the 3 groups. Ewes in the Group 1 set in estrus later (p<0.05; $50.0{\pm}7.29h$) than the ewes in Group 2 ($24.5{\pm}3.58$) and 3 ($32.5{\pm}3.58h$). The duration of estrus, ovarian size and ovarian response (number of CL and LF) did not differ significantly (p>0.05) among the 3 groups. The proportion of ewes with a superovulatory response (${\geq}2$ CL) was the lowest (50%) in Group 1 treated with FSH alone but ova/embryo recovery (100%) and fertilization (100%) was significantly (p<0.05) higher than Group 2 (58.3 and 85.7%, respectively) and Group 3 (48.6 and 50%, respectively). It is concluded that in tropical fine wool sheep, there is no difference in the 3 treatments for yield of good quality embryos but ovarian response and ovulation rate increased on additional use of PMSG and GnRH respectively to FSH alone.

• Reproductive and Developmental Biology
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• v.39 no.2
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• pp.29-36
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• 2015

Pigs are considered an ideal source of human disease model due to their physiological similarities to humans. However, the low efficiency of in vitro embryo production (IVP) is still a major barrier in the production of pig offspring with gene manipulation. Despite ongoing advances in the associated technologies, the developmental capacity of IVP pig embryos is still lower than that of their in vivo counterparts, as well as IVP embryos of other species (e.g., cattle and mice). The efficiency of IVP can be influenced by many factors that affect various critical steps in the process. The previous relevant reviews have focused on the in vitro maturation system, in vitro culture conditions, in vitro fertilization medium, issues with polyspermy, the utilized technologies, etc. In this review, we concentrate on factors that have not been fully detailed in prior reviews, such as the oocyte morphology, oocyte recovery methods, denuding procedures, first polar body morphology and embryo quality.

• Journal of Embryo Transfer
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• v.30 no.3
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• pp.149-159
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• 2015

This study was designed to know the possibility in repeat uses of elite donor cows for getting mass production of OPU-derived embryo production (OPU-IVP). Ultrasound transvaginal ovum pick-up (OPU) performed in 6 Korean native cows was aged 4 to 10 years old. The aspiration of immature oocytes for OPU derived embryo was carried out 2 times per week, and OPU-IVP of $1^{st}$ period was carried out 22~48 sessions from each donors. And the break time for OPU-IVP of $2^{nd}$ period after $1^{st}$ OPU from each donors were 2~25 months. The OPU-IVP of $2^{nd}$ period each donors conducted total 15~65 times for 2~8 months by an ultrasonographic, was guided follicular aspiration system. The average numbers of collected oocytes, grade 1 + grade 2(G1+G2) oocytes and cleavage embryo from $1^{st}$ period OPU-IVP were significantly differences between donors (p<0.05). Total collected oocytes of donor D were significantly higher compared with donors of A, B, C, E and F (average 17.0 per session vs. 11.2, 10.1, 8.5, 10.2 and 9.6; p<0.05) and also oocytes of G1+G2 were significantly higher compared with r A and D and subsequently to donors of B, C, E and F (average 7.9 and 8.5 per session vs. 5.0, 2.7, 6.0 and 1.6; p<0.05). Cleavage rate of donor D was significantly higher compared with donors of A, B, C, E and F (average 13.1 per session vs. 10.1, 9.1, 6.9, 8.9 and 6.7; p<0.05). The average numbers of OPU-IVP for $1^{st}$ period was significantly higher from donors of B, D and E than those from donors of A, C and F (average 6.5, 7.1 and 6.5 per session vs. 3.5, 4.2 and 2.8; p<0.05). The possibility investigation of $2^{nd}$ OPU-IVP was carried out after 2~25 months rest periods from $1^{st}$ period OPU session. Total average numbers of collected oocytes, cleavages and blastocyst development rates were significantly higher from $1^{st}$ period OPU compared with $2^{nd}$ period one (p<0.05). The OPU-IVP efficiency by break for more embryo production from elite cow was analysis comparing without rest of donor A, under 6 months rest period as B and over 6 months rest period as C and then the average numbers of collected oocytes, cleavages and blastocysts were significantly higher from A group (11.8, 9.5 and 5.2 per session) than those from B and C groups (7.9, 6.2 and 2.6 vs. 9.2, 7.5 and 3.9, p<0.05), and also C group was significantly higher than B group. In conclusion, $1^{st}$ period OPU-IVP was more efficient compared with $2^{nd}$ period repeated uses of donor, and the break times for additional production of embryo on donor were needed more than over 6 months after $1^{st}$ period OPU-IVP. This repeating uses of elite donor cows given more emphasis for getting the opportunity on mass production of elite cow OPU-IVP embryo should be increased G1+G2 possibility of genetic improvement of livestock within short period.

• Journal of Embryo Transfer
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• v.31 no.3
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• pp.191-197
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• 2016

Historically, Korea old cattle had been consisted with various lines of coat color brindle, black and white-brown breeds or more. The two rare lines of black and white coat color are maintained for animal resources and preserved critically. The present study was carried out to evaluate potential usage of cysteamine supplementation during in vitro matration (IVM) and in vitro culture/production of embryo (IVP) by transvaginal ultrasound-guided follicle aspiration (Ovum Pick-Up: OPU) for the establishment of cryo-banking system. Immature slaughterhouse-derived cumulus-oocyte complexes (SL-COCs) were matured in IVM medium supplemented with 0, 0.1, 0.3 or 0.9 mM cysteamine, and then cultured in mSOF-BAS for 8 days after in vitro fertilization. The treatment of 0.1 mM cysteamine on SL-COCs showed higher rate of blastocyst, so OPU-derived COCs from rare breeds were matured in TCM media supplemented with or without 0.1 mM cysteamine, FSH and 5% FBS. The embryos were evaluated their developmental stages on day 8. During IVM, cysteamine treatment significantly increased the embryo production rate of slaughterhouse-derived COCs (19.6% vs. 30.5%). The presence of cysteamine during IVM of OPU-derived COCs from rare Korean cattle breeds (albino white and black line) also increased embryo production rates than those from SL-COCs (27.4% vs. 41.9% and 36.4%). With these results, cysteamine treatment during IVM is one of key factors IVP of blastocysts to establish banking system of endangered rare Koarean cattle with OPU derived transferable blastocysts.

• Journal of Animal Reproduction and Biotechnology
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• v.34 no.4
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• pp.305-310
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• 2019

Embryos produced with serum show the alterations in their ultrastructure, impaired compaction, abnormal blastulation, aberrant mRNA expression profiles and large calf syndrome with greater incidences of stillbirths and deaths after birth. The aim of the present study was to describe in vitro embryo production by analyzing embryo production, fetal production and pregnancy rate in free-serum medium. The OPU-IVP data used in this study from 2016. Approximately, sixteen cows (Hanwoo), which belonged to the Institute of Gyeongsang National University, were used. Two experimental group is used in this study. Serum groups were conducted in March to July and free-serum group was conducted in September to December. The recovered cumulus-oocyte complexes were morphologically classified to four grades based on the compaction of cumulus cells layers and homogeneity of the cytoplasm. The number of oocyte was significantly greater in serum groups than that in free-serum groups (29.61 ± 0.63 vs. 15.6 ± 0.62; p < 0.05). Between serum and free-serum groups indicate that average of 1st and 2nd grade oocytes were no difference (2.38 ± 1.67 vs. 2.38 ± 1.48; p > 0.05), but number of 3rd and 4th grade oocytes were greater in serum groups than that in free-serum groups (7.31 ± 7.64 vs. 5.60 ± 6.29; p < 0.05). Embryo cleaved competence was higher in rate in free-serum groups than that in serum groups (62.1% vs. 58.3; p < 0.05). However, blastocyst developmental rate was no difference between serum and free-serum groups (33.1% vs. 43.5%; p < 0.05). 986 recipients were used for embryo transfer. Pregnancy rate was indicated that between serum and free-serum group was no difference (54.6% vs. 56.3%; p < 0.05). In conclusion, we developed the free-serum system for production of in vitro bovine embryos in order to meet the developmental and qualitative requirements for large scale commercial use.

• Journal of Embryo Transfer
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• v.23 no.3
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• pp.167-175
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• 2008

This study was performed to investigate the result that in-vivo or in-vitro embryos of Hanwoo cows were transferred to Holstein cows. Seventeen Hanwoo cows were used as donors for production of in-vivo embryos and fresh hanwoo in-vivo embryos were transferred to 1,150 Holsteins. And 2 embryos were transferred to 188 Holstein recipients to produce twin calves. Diagnosis on pregnancy was performed by rectal palpation at $60\sim90$ days after transfer. The pregnancy rate of Holstein recipients was 55.8% after transfer with Hanwoo in-vivo embryos and 38.2% after transfer with Hanwoo in-vitro embryos. The delivery rate of pregnant Holstein recipients was 88.4% after transfer with Hanwoo in-vivo embryos and 75.6% after transfer with Hanwoo in-vitro embryos. The rate of delivery of Holstein recipients transferred with two Hanwoo embryos was 36.2% and the rate of twin production was 25.9%. The rate of twin production by embryo transfer with in-vivo embryos was 30.4%, whereas the fate with in-vitro embryos was 15.6%. The pregnancy rate according to the grade of corpus luteum of Holstein recipients transferred with Hanwoo in-vitro embryos was 41.5 and 36.0% for A and B grade, respectively. The pregnancy rate according to the transfer in site in the uterine lumen of recipients was 40.9 and 32.7% for anterior and middle site, respectively. The pregnancy rate according to day of embryo transfer after estrus of recipients was 45.5, 38.8 and 39.7% for day 6, day 7 and day 8, respectively. There was difference of pregnancy rate according embryo transfer technician ($30.5\sim45.8%$) individual dairy farm ($21.1\sim51.0%$). These results are supposed to indicate that the rate of pregnancy after transfer with Hanwoo embryos to Holstein recipients was similar to that within the same breed, and consequently that this method would be beneficial to enhance the productivity in Hanwoo reproduction.

• 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 Embryo Transfer
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• v.33 no.3
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• pp.139-147
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• 2018

Somatic cell nuclear transfer (SCNT) is a useful biotechnological tool for animal cloning. Until now, SCNT has been inefficient, especially in dog. It is believed that an embryo developmental block in SCNT embryos is cause of low production efficiency. However, no studies have been performed on canines for embryo developmental block. In this study, we attempted to evaluate the beneficial role of EDTA in canine parthenogenic (PA) embryos development to overcome embryo developmental block. The PA embryos were divided into 0.01 mM EDTA treated and non-treated groups. Embryo developmental efficiency was measured by activating chemically parthenote. After EDTA induction, PA embryos were evaluated for embryonic development, Reactive Oxygen Species (ROS) activity, mitochondrial integrity, ATP production and genomic activation. The EDTA treated PA embryos showed significantly higher survival rate and improved cavity formation compared to non-treated. Furthermore, cytoplasmic ROS level was mitigated and mitochondrial membrane potential was found significantly higher in EDTA treated group followed by higher ATP production. Moreover, major embryonic genomic activation specific markers/factors were also elevated in EDTA treated group. Conclusively, we elucidated that EDTA showed substantially positive effect to overcome embryo developmental block in canine.

• Journal of Embryo Transfer
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• v.28 no.1
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• pp.7-12
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• 2013

Body condition score (BCS) is a useful management tool for distinguishing differences in nutritional needs of cows in the herd. Although it is not always possible to quantify the nutrient content of the feed supplied to the donor cow, the nutritional status can be determined by the BCS. The objective of this study was to evaluate in vivo embryo production, return to estrous of donor and pregnancy rate of recipients following BCS in Hanwoo superovulation. Sixty nine Hanwoo donor cows were flushed on day 7 of estrus cycle with same FSH and artificial insemination by the same technicians. Embryos were recovered on 7 days after the third insemination by flushing the uterus with Embryo Collection Medium. The results obtained were as follows: No differences were observed in the efficiency of superovulation rates regardless of BCS ($${\leq_-}2.0$$, 2.5 to 3.0, and $${\geq_-}3.5$$). The mean number of total embryos was each $5.20{\pm}0.86$, $11.56{\pm}1.04$, and $6.23{\pm}1.07$. The mean number of transferable embryo from $${\leq_-}2.0$$, 2.5 to 3.0, and $${\geq_-}3.5$$ of BCS was $2.60{\pm}0.87$, $7.94{\pm}0.89$, and $4.75{\pm}1.32$, respectively (p<0.05). Return to estrous regardless of donor BCS was no difference. The pregnancy rates of recipient were BCS $${\leq_-}2.0$$ 11.76%, 2.5 to 3.0 40.79%, and $${\geq_-}3.5$$ 11.11%, following transfer of fresh embryos produced in vivo, respectively. These results indicate that if the Hanwoo with BCS 2.5 to 3.3 are used for donor and recipient, the embryo production and the conception rate will be greater.