• Asian-Australasian Journal of Animal Sciences
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• v.20 no.2
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• pp.194-199
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• 2007

Effects of recombinant bovine somatotropin (bST) on plasma hormonal concentration, embryo quality, and pregnancy rate were examined during the superovulation and synchronization treatment in donor and recipient cows. Hanwoos (Korean native beef cattle) were treated with controlled internal drug release (CIDR) combined with bST (CIDR+bST) or without bST (CIDR) as donor cows. The embryos recovered from donors were transferred into Holstein recipient heifers treated with bST (CIDR+bST) or without bST (CIDR) for synchronization. The correlation between IGF-I and P4 showed a positive pattern in the CIDR+bST group (r=0.44, p<0.01), but a negative pattern was shown in the CIDR group (r = -0.59, p<0.02) at day 7 of estrous cycles. Although the number of recovered, transferable, and degenerated embryos was not different, quantities of grade 1 (excellent) embryos in CIDR+bST group were significantly higher than those of the CIDR group (p<0.01). The pregnancy rate was higher in the CIDR+bST recipient group compared to CIDR group (p<0.05), when the embryos were recovered from the donors treated with CIDR. However, the pregnancy was maintained highly in both recipient groups, when the embryos were produced by CIDR+bST treated donors. It can be concluded that bST administration combined with CIDR is an effective method for superovulation and synchronization treatment to stabilize plasma hormonal levels, to obtain excellent quality of embryos, and to get higher pregnancy rate.

• 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.

• Journal of Embryo Transfer
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• v.14 no.2
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• pp.113-119
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• 1999

This study was performed to improve the development of the in vitro fertilized bovine embryos by the condition of in vitro maturation. COCs were matured in TCM 199 supplemented with 0.1% PVA, 10ng/ml EGF, Hormones (5$\mu\textrm{g}$/ml FSH, 10 IU hCG, 1 $\mu\textrm{g}$/ml estradiol 17-$\beta$) or granulsa cell+Hormones atmosphere 39$^{\circ}C$, 5% CO2, 95% air for 24hrs. Matured oocytes were fertilized with frozen-thawed semen capacitated with 5mM caffein in BO medium for 20 hrs. IVF embryos were cultured in TCM 199 containing with hormones(same as matured medium), 10% FBS and co-culture with bovine oviduct epitherial cells. Maturation rates of COCs were showed 73.8%, 78.5%, 83.2% and 87.6% respectively, and were significant differences between PVA, EGF, and Hormones, GC+Hormones(p<0.05). The cleavage rates of IVF embryos were revealed 72.5%, 78.4%, 82.3% and 84.2% and showed same tendency as maturation rates(p<0.05). The blastocysts matured by above maturation condition and cultured for 7~10 days after fertilization had 34.4, 43.6, 52.3 and 59.3 cells had no differences among the treatments. These results suggest that high molecules as a substitutes of serum and growth factor may induce nuclear resumption of COCs but we need more study to produce transferable IVF blastocysts by use of that agents.

• Journal of Embryo Transfer
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• v.16 no.2
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• pp.99-106
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• 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
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• v.20 no.3
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• pp.311-315
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• 2005

This study was conducted to investigate the effects of disregarding of heat detection on embryo production in superovulation of Hanwoo cows. Donors which showed 1 or 2 times of normal heat and had no abnormality of reproductive tract were selected The superovulation was performed injection of 2.5 mL FSH (Antorin R-10, Japan) 2 times on 5 days before next heat and continuously with the reduction of dose to 0.5 mL of first injection 2 times in a day for next 3 days. Otherwise, the donors of CIDR group were inserted CIDR plus (with the capsule of estradiol benzoate 10 mg) on Day 10 from standing heat for 9 days. On 6 days from insertion of CIDR, FSH was injected above same manners. The response according to the natural heat and CIDR were $82.2\%,\;89.7\%$, respectively. There were no difference between both treatments. The recovery rates of embryos were 7.7, 10.5, respectively and transferable embryos were 3.4, 3.8. There showed significant difference between both treatments (p<0.05). These results suggested that disregarding of heat detection in superovulation could be produced transferable embryos for embryo transfer and preserve the donors from the excess hormonal administration and maintain the economical lift span of genetically available Hanwoo donors.

• Journal of Embryo Transfer
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• v.21 no.2
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• pp.147-156
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• 2006

The objective of this study was to supply excellent genetic resources to livestock farms by transferring embryos produced by genetically superior Korean cows (Hanwoo). Eighty Hanwoo donors were superovulated with gonadotropin ($Folltrpin^(R)\;or\;Antorin^(R)$) for 4 days combined with or without progesterone releasing intravaginal device (CIDR) insertion. The collected fresh or frozen-thawed embryos were transferred to 226 farm recipients. In this study, the effect of CIDR insertion in combination with gonadotropin ($Folltrpin^(R)$) treatments initiated at the random stage of estrous cycle on embryo production was evaluated and compared to conventional superovulation protocol. Moreover, the effect of gonadotropin ($Antorin^(R)$) dose in CIDR-treated Hanwoo donors on the embryo yield was determined. In addition, the effects of embryos (fresh vs. frozen-thawed), embryo transfer person, seasons and farms on the pregnancy rate were evaluated. In Hanwoo donors, CIDR insertion in combination with $Folltrpin^(R)$ treatments regardless of estrous detection resulted in increased numbers of total ova (6.5 vs. 5.8) and transferable embryos (3.9 vs. 3.2) compared to the conventional superovulation protocol (p<0.01). In CIDR-treated Hanwoo donors, the higher dose of $Antorin^(R)$ (36 vs. 28 mg) resulted in the increased number of transferable embryos (8.3 vs. 5.4, p<0.05). The embryos (fresh 43.9% vs. frozen-thawed 23.1%) and embryo transfer person (53.9 vs. $0{\sim}16.7%$) significantly affected the pregnancy rate after embryo transfer (p<0.01). These results suggest that CIDR-based superovulation protocol may be effectively used for production of superior Hanwoo embryos and, multiple ovulation and embryo transfer in Hanwoo might be effectively applied for livestock improvement if pregmancy rate with frozen-thawed embryos and embryo transfer skill would be improved.

• Journal of Practical Agriculture & Fisheries Research
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• v.16 no.1
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• pp.115-122
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• 2014

Korean native cattles (Han woo) are divided yellow, tiger(blinder), and black by coat color. Chikso has tiger coat color, which cultured about 1,700 heads within Korea. However there were a few researches reported about preservation and production. This study was conducted to investigated that the effect of follicle stimulating hormone (FSH) dosage(240 mg and 280 mg) on the embryo production, pregnancy and calf birth weights. There were no difference total embryo and transferable embryo between FSH 240 mg or 280 mg treatment, but pregnancy rate was increased 46% in embryos derived from 280 mg group than 240 mg group's 29.6%. The birth weight of Chikso calves was similar between artificial insemination(25.6 kg) and embryo transfer(24.8 kg) groups.

• Journal of Embryo Transfer
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• v.20 no.2
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• pp.185-190
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• 2005

This study was performed to investigate the effects of multiple superovulation and parity on embryo production in Hanwoo cows. Donors were superovulate 4 times $1\~2$ months interval and inserted CIDR plus (with the capsule of estradiol benzoate 10mg) on Day 10 from standing heat for 9 days and injected 2.5ml FSH (Antorin R-10) 2 times in a day on 6th day to 10th day from insertion of CIDR and the doses of FSH were decreasing 0.5ml on every 2 times. On 3th day of FSH injection, 25ml $PGF_2{\alpha}$ were injected i.m. and on 4th day, CIDR was removed. After 2 days from removing CIDR, AI was performed 2 times 12 hour apart with 2 straws of Korean Proved frozen Semen and simultaneously 200ug/ml GnRH was injected and embryos were recovered on 7th day from Al. The response rates of superovulated donors were $85.7\%,\;90.5\%,\;62.5\%,\;100\%$ from 1 to 4 times of superovulation, respectively. There were significant differences among No. of superovulation times (P<0.05). The results of transferable embryos were 3.7, 3.4, 3.4, 5.7 from 1 to 4 times of superovualtion, respectively. There were no differences among No. of superovulation times. The results of transferable embryos were 2.5, 3.0, 5.3, 3.0, 3.4 form heifer, first born to 4 the born, respectively. There were significant differences among the parities of donors (P<0.05). These results suggested that even 4 times of superovulations of Hanwoo donors could be able to recover transferable embryos, it might be used the donors maximally and improved the adaptation of embryo transfer to farms safely.

• Korean Journal of Veterinary Research
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• v.33 no.4
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• pp.757-762
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• 1993

The ovaries of Korean Native cows or heifers were obtained from an abattoir and kept on 20 to $25^{\circ}C$ and transported to laboratory within 2 hrs. The follicular oocystes were collected from 2~6mm follicles in diameter and classified into 3 grades by the morphology of cumulus cells attached. The oocytes were matured in vitro(IVM) for 24 hrs. in TCM-199 supplemented with $23{\mu}g/ml$ FSH, $10{\mu}g/ml$ LH, $1{\mu}g/ml$ estradio-17 ${\beta}$ and granulosa cells at $39^{\circ}C$ under 5% $CO_2$ in air. They were fertilized in vitro(IVF) by incubation for 12 hrs. of epididymal spermatozoa pretreated with heparin, and then the zygotes were co-cultured in vitro(IVC) with oviductal epithelial cells for 7 to 9 days. Assessment of maturation revealed that 93.0%(147/158) of grade I oocytes had expanded of cumulus cells, which was higher(p<0.05) than the 79.4%(85/107) of grade II oocytes. Compared to epididymal sperm(32.9%), the insemination with frozen and thawed sperm resulted in slightly lower(20.5%), but not significant, development to morulae and blastocysts from grade I oocytes. Co-culture of bovine IVF embryos with oviductal epithelial cells improved the development to transferable embryos significantly(38.1%), compared to co-culture with granulosa cells(20.0%). When VF bovine embryos were vitrified at blastocyst, the post-thaw survival rate was obtained higher resulf for 1 min. equilibration time(82.6%) or 2 min.(73.9%) than 3 min.(18.2%) in EFS solution.

• Reproductive and Developmental Biology
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• v.35 no.3
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• pp.233-238
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• 2011

This study was performed to investigate the FSH levels for superovulation procedure in Korean Native Cattle (Hanwoo). The effectiveness of 200 mg and 400 mg of FSH to initiate superovulation was examined in Hanwoo. Donors, at random stages of the estrous cycle, received a CIDR 7 days later, 200 mg FSH group was treated with 40, 30, 20, 10 mg FSH levels in declining doses twice daily by intramuscular injection fur 4 days. Also, 400 mg FSH group was treated with 80, 60, 40, 20 mg FSH levels. On the 3rd day administration of FSH, 25 mg $PGF_2$ ${\alpha}$ was administered and CIDR was withdrawn. Donors were artificially inseminated twice at 12 hr intervals. The donor cattle received 250 ${\mu}g$ GnRH at time of 1st insemination and embryos were recovered 8 days after the 1st insemination. As a results, average number of CL treated with FSH 200 mg was higher as $20.9{\pm}1.20$ than $15.8{\pm}0.63$ for donors treated with FSH 400 mg, respectively(p<0.05). Treated group of 200 mg FSH level increased (p<0.05) the number of embryos recovered per procedure compared to 400 mg FSH level ($18.2{\pm}1.18$ vs. $12.38{\pm}0.52$, respectively). When treatment of 200 mg FSH was performed, average transferable embryos/ova increased (p<0.05) to $14.1{\pm}1.12$ from $6.8{\pm}0.33$ of treated of 400 mg FSH. Group of 200 mg FSH increased (p<0.05) to $8.3{\pm}0.76$ from $2.0{\pm}0.26$ in morula stage compare to 400 mg FSH group. Mean of total early blastocyst and expanded blastocyst stage embryos was similar (p<0.05) between the 200 mg and 400 mg FSH levels group ($4.7{\pm}1.19$ vs. $2.9{\pm}0.18$ and $1.2{\pm}0.40$ vs. $1.9{\pm}0.17$). These results suggest that 200 mg FSH level-based superovulation protocol with CIDR may be effectively used fur production of superior embryos in Hanwoo. In other words, the less level of FSH may be effectively applied for Hanwoo (Korean Native Cattle), because Hanwoo was smaller body size than beef or daily cow.