• 한국수정란이식학회지
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• 제9권3호
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• pp.221-227
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• 1994

This study was conducted to examine the condition of in vitro culture system and the viability after embryo transfer of in vitro matured-in vitro fertilized (IVM-IVF) bovine embryos. The in vitro development to the blastocyst stage was enhanced by supplying bovine serum albumin(BSA) to co-culture medium with bovine oviduct epithelial tissue(BOET) compared with that in medium supplemented with fetal bovine serum(FBS) (41.2% vs. 26. 3%, P<0.05). After transfer of IVM-IVF blastocysts into the uterine horn of recipient females (Aberdeen Angus), one was pregnant to term and produced a head of male Korean native calf. These results confirm that the in vitro development of IVM-IVF bovine embryos is affected with different protein source in co-culture with BOET, and IVM-IVF embryos can develop to term after in vitro culture and embryo transfer.

• Asian-Australasian Journal of Animal Sciences
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• 제14권9호
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• pp.1342-1351
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• 2001

Over the years, the embryo transfer industry has grown from the simple collection & transfer of embryos into an advanced field of embryo biotechnology. Currently a large demand exists for bovine oocytes and early embryos in both research and commercial settings. Bovine embryos can now be produced in-vitro. Primary oocytes collected from antral follicles of abattoir - obtained ovaries can be induced to undergo the maturation process. In-vitor maturation system, however must ensure that the resulting oocyte is capable of undergoing normal fertilization and yields a zygote competent of developing to term after embryo transfer. Sperm preparation for IVF has improved with the use of heparine. The use of co-culture system has proved beneficial in circumventing the developmental block in IVM/IVF bovine embryos.

• 한국수정란이식학회지
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• 제9권3호
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• pp.229-234
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• 1994

This experiment was carried out to develop the model system for mass production of biomedical and nutritional proteins (human proteins) through mamraary gland of the transgenic cattle produced by gene manipulation and embryological technologies. Human growth hormone gene fused with rat $\beta$-casein gene promoter was microinjected into pronuclei of one cell bovine embryos produced by in vitro fertilization. After microinjection, embryos were cultured in vitro for 6 or 7 days. Twenty embryos reaching to blastocysts were transferred to 10 beef recipients, each receiving two embryos. Recipients were diagnosed for pregnancy by rectal palpation at 76 days after embryo transfer. One of them was pregnant to term and produced a female calf weighing 21 kg at 280 days following embryo transfer. DNA was extracted from umbilical cord tissue and blood of calf born for confirming gene insertion. As determined by Southern hybridization, the transgene was not found.

• 한국수정란이식학회지
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• 제10권3호
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• pp.229-236
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• 1995

The experiments reported here take advantage of the large number of in vitro matured and in vitro fertilized(IVM /IVF) bovine oocytes which can be produced, permitting the design of controlled experiments to establish a simple defined medium for the study of early embryo requirements. A total of 1,386 IVM /IVF oocytes were used to compare a simple defined medium(KSOM) with more complex culture conditions used successfully for culture of bovine embryos but do not permit study of specific requirements. All experiments were extensively replicated factorials. In Experiment 1, KSOM was superior to Menezo B$_2$ medium in producing morulae plus blastocysts from IVM /IVF oocytes(33 vs 20%, P<0.()5). The yield of morulae plus blastocysts with KSOM was 22% and with BRLC added was 30%. In Experiment 2, (a 2x2 factorial of KSOM with or without BRLC and 0, 1 ng /ml of platelet derived growth factor, PDGF) more morulae plus blastocysts (40%) were produced in KSOM-BRLC co-culture containing 1 ng /ml PDGF than in the control KSOM(12%). In Experiment 3, there was no dose response when 0, 1 and 5 ng /ml of PDGF were added. The results with simple defined KSOM medium are sufficiently promising to indicate that specific requirements of the embryo may be examined in future studies with KSOM as a base.

• 한국수정란이식학회지
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• 제24권4호
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• pp.253-257
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• 2009

To enhance the embryo preservation technology and better application of embryo transfer technique to the field (dairy science or animal reproduction. etc.), we examined the viabilities of bovine embryos produced in vitro and in vivo after cryopreservation according to their developmental stage and thawing temperature. Bovine embryos from in vivo/vitro fertilization (Hanwoo) were examined at day 7, 8, and 9. Survival rates and total cell numbers of in vivo fertilized embryos were as follows: morulae 68.8% and $67\;{\pm}\;6.0$; blastocysts 80.5% and $120\;{\pm}\;10$; expanded blastocysts 77.4% and $138\;{\pm}\;9.7$, respectively. Rates of embryo development for blastocysts and expanded blastocysts after thawing were significantly higher than that of morula stage embryos (p<0.05). While survival rates of in vitro fertilized embryos according to developmental stage showed no significant difference among groups (morula 67.9%; blastocyst 74.3%; and expanded blastocyst 79.4%), total cell numbers were significantly lower than those of other groups (morula $64\;{\pm}\;5.9$; blastocyst $116\;{\pm}\;8.7$; and expanded blastocyst $135\;{\pm}\;9.1$) For the viability according to thawing temperature, survival rate was higher in $37^{\circ}C$.

• 한국수정란이식학회지
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• 제14권3호
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• pp.171-176
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• 1999

This study was carried out to investigate the effect of co-culture system(bovine oviduct epithelial cells; BOEC) and defined culture system (modified TALP ; mTALP) on the conception of embryos transferred, and pregnancy and twin birth rates after transfer of fresh or frozen-thawed bovine blastocysts produced in vitro were also evaluated. Oocytes from the slaughterhouse ovaries were matured and fertilized using general protocol. The results obtained were as the following. The pregnancy rate after transfer was higher in co-culture group than in mTALP group, but was not signficantly different, and there is no difference between fresh embryo group and frozen-thawed embryo group in conception rate. The conception rate was not different whether 3∼4 blastocysts or 2 blatocysts transferred into a recipient, but the production rate of twin calves was significantly higher (p<0.05) when 3∼4 embryos transferred. The average birth weight of twin calves(24.38kg) was numerically, but not significantly lighter than that of single calves(26.68kg).

• 한국가축번식학회지
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• 제18권4호
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• pp.251-256
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• 1995

Two experiments were conducted to study the production of in vitro fertilized bovine embryos and the viability of blastocysts cryopreserved by vitrification. In experiment 1, production rate of in vitro matured bovine oocytes after fertilization in medium containing bovine oviduct epithelial cells (BOEC), cumulus cells and granulosa cells to blastocysts were 18.4, 14.6 and 13.1%, respectively. Developmental percentages of blastocysts produced at day 6, 7 and 8 were 8.5, 10.6 and 15.2% respectively. Hatching rate of bovine embryos produced was 60.0%. In experiment 2, post-thawed surviving embryos in a vitrification solution consisting of 7.15M ethylene glycol, 2.5 mM ficoll and 0.3 M sucrose were 36.4% (56/154). Also, survival rate of bovine embryos after exposed to vitrification solution at 1, 2, 3, 4 and 5 min were 84.0, 88.0, 71.0, 48.0 and 24.0% respectively.

• 한국수정란이식학회지
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• 제10권2호
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• pp.171-175
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• 1995

This study was performed to establish the condition and the methods for the techniques of insertion the isolated blastomere cells into cytoplasm, in order to research the develop-mental ability of bovine embryo blastomere cells in vitro produced. After 24h in vitro ovary maturation with the ovaries from a slaughter house, in vitro fertilization was performed to the vital sperms which their mobility were decided by percoll gradient method, with 2~8 cell stage embryos, the blastomeres were isolated in $Ca^2$+. $Mg^2$+-free PBS, and following that embedded into agar and alginate solution, respectively. The rates of in vitro develop-ment are as follows ; in agar embedded 11 among 120(9.2%) 1 /2~1 /3 blastomers cleaved and 6 among 93(6.5%) 1 /4~1 /8 blastomeres cleaved. In sodium alginate-embedded 14 among 84(16.7%) 1 /2~1 /3 blastomeres cleaved and 6 among 85(7.1%) 1 /4~1 /8 blastomeres cleaved. In case of Na-alginate, the rate of the cells were better than those of agar. The results suggest that the techniques for embeeding the isolated blastomeres into gel may help cloning of bovine early embryo without nuclear transplantation.

• 한국수정란이식학회지
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• 제33권3호
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• pp.169-175
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• 2018

It has been claimed that artificial insemination (AI) of cows with frozen-thawed semen treated with commercially produced kits, Wholemom (in favour of female gender) increases the birth chance of calves with desired sex ratio by approximately 85% without decrease of pregnancy rates. Hence, this study was conducted to investigate the efficacy of wholemom kits as combined with frozen-thawed bovine semen during in vitro fertilization on the in vitro fertilization and developmental efficiency and sex ratios such as some reproductive parameters in bovine. For this, 1,737 oocytes were in vitro fertilized and developed. Agglutination effects on bovine after treatment of Wholemom kit were observed by time passage and dose respectively. To determine sex of embryos, Bovine embryo Y-specific gene primers(ConEY) and Bovine specific universal primer(ConBV) were used as multiple PCR method. Fertilization rate of wholemom-treated group was significantly lower than its of control group[66.9% (1,156/1,737) in Wholemom-treated group; 75.0% (610/813) in control group]. However, developmental rate after fertilization of both wholemom-treated and control groups were not significantly different [26.1% (404/1,156) in Wholemom-treated group; 27.4% (224/610) in control group]. Sex ratio of in vitro fertilized embryo with frozen-thawed semen treated with wholemom kit was determined by multi PCR. Female ratio in wholemom-treated group [85.4% (173/201)] was significantly higher than its of control group [47.2% (66/141)]. In conclusion, wholemom treatments of semen used in the in vitro fertilization and development of bovine oocytes provided increase in female ratio with decrease of fertilization rate.

• 한국수정란이식학회지
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• 제8권2호
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• pp.143-149
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• 1993

The objective of this study was to produce calves derived from in vitro fertilization of in vitro matured follicular oocytes. Oocytes aspirated from small antral folicles of ovaries obtained at a local slaughter house were matured and fertilized in vitro. At l8hrs after insemination with Korean native cattle semen, oocytes were co-cultured for 6~7 days by utilizing co-culture system with bovine oviduct epithelial cell. After co-culture, good or excellent quality late morulae or early blastocysts were selected by morphological criteria under stereo microscope. Selected embryos were transferred to recipients on day 6 or 7 (estrus = day 0). Recipients were monitored by observation for estrus and rectal palpation after 60 days from embryo transfer. One of them went to term with the birth of a calf. This case is the first production of calf derived from in vitro fertilization in Korea.