• Journal of Embryo Transfer
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• v.11 no.2
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• pp.137-144
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• 1996

The present study was carried out to obtain the pregnancy and delivery rate following transfer of fresh and frozen-thawed Korean native cattle(KNC) blastocysts(1~4 em-bryos / head) produced in vitro to Holstein recipients. The pregnancy rate of fresh and frozen-thawed KNC blastocysts produced in vitro was 50%(7 /14 heads) and 38.5%(5 /13 heads), respectively. The pregnancy rate of frozen-thawed KNC blastocysts produced in vitro frozen using 1.5M ethylene glycol and 1.4M glycerol for cryoprotectant was 33.3%(2 /6 heads) and 42.9 %(3 /7 heads), respectively. Seven calves including 2 sets of twin were born fiom 5 pregnant recipients receiving eleven fresh blastocysts. Three pregnant recipients were aborted among four pregnant recipients receiving twelve frozen-thawed blastocysts and one calf was born from the rest of one pregnant recipient.

• Asian-Australasian Journal of Animal Sciences
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• v.33 no.7
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• pp.1087-1095
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• 2020

Objective: The present study aimed to evaluate the effects of γ-aminobutyric acid (GABA)-producing bacteria (GPB) on in vitro rumen fermentation and on the growth performance and meat quality of Hanwoo steers. Methods: The effects of GPB (Lactobacillus brevis YM 3-30)-produced and commercially available GABA were investigated using in vitro rumen fermentation. Using soybean meal as a substrate, either GPB-produced or commercially available GABA were added to the in vitro rumen fermentation bottles, as follows: control, no additive; T1, 2 g/L GPB; T2, 5 g/L GPB; T3, 2 g/L autoclaved GPB; T4, 5 g/L autoclaved GPB; T5, 2 g/L GABA; and T6, 5 g/L GABA. In addition, 27 Hanwoo steers (602.06±10.13 kg) were subjected to a 129-day feeding trial, during which they were fed daily with a commercially available total mixed ration that was supplemented with different amounts of GPB-produced GABA (control, no additive; T1, 2 g/L GPB; T2, 5 g/L GPB). The degree of marbling was assessed using the nine-point beef marbling standard while endotoxin was analyzed using a Chromo-Limulus amebocyte lysate test. Results: In regard to in vitro rumen fermentation, the addition of GPB-produced GABA failed to significantly affect pH or total gas production but did increase the ammonia nitrogen (NH₃-N) concentration (p<0.05) and reduce total biogenic amines (p<0.05). Animals fed the GPB-produced GABA diet exhibited significantly lower levels of blood endotoxins than control animals and yielded comparable average daily gain, feed conversion ratio, and beef marbling scores. Conclusion: The addition of GPB improved in vitro fermentation by reducing biogenic amine production and by increasing both antioxidant activity and NH₃-N production. Moreover, it also reduced the blood endotoxin levels of Hanwoo steers.

• Biotechnology and Bioprocess Engineering:BBE
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• v.9 no.6
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• pp.440-446
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• 2004

Interest in in vitro study of entomopathogenic fungi, including Cordyceps species, has been increasing due to their valuable bioactive compounds and biocontrol effects. Among Cordyceps species, in vitro stromata of C militaris has been successfully produced and cultivated for industrial purposes. However, genetic study on in vitro stromata formation of C militaris has not been carried out yet. Here, relationship between mating system and perithecial stromata formation of C militaris is reported. Mating system was determined by observing perithecial stromata formation from mono-ascospore cultures and their pair-wise combinations. Certain combinations of mono-ascospore strains produced perithecial club-shaped stromata, whereas other combinations produced either no stromata or only abnormal non-perithecial stromata. Similarly, mono­ascospore cultures without combination produced either no stromata or only abnormal non­perithecial stromata. Despite obvious heterothallism, self-fertility was occasionally observed in few strains of C militaris. These observations indicated that C militaris behaves as a bipolar het­erothallic fungus and requires two mating compatible strains in order to produce regular club­shaped perithecial stromata, a fundamental requirement for its industrial cultivation.

• Journal of Plant Biotechnology
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• v.6 no.3
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• pp.181-188
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• 2004

Zingiber officinale buds from the rhizomes were used to produce in vitro shoots. These explants produced the largest number of multiple shoots, 9.8 shoots per explant, when were cultured on MS (Murashige and Skoog 1962) medium supplemented with 2.0 mg/L benzyladenine (BA) and 2.0 mg/L indole butyric acid (IBA). This medium was also found to be suitable for in vitro propagation of other Zingiberaceae species: Alpinia conchigera, Alpinia galanga, Curcuma domestica, C. zedoaria and Kaempferia galanga. Both C. domestica and C. zedoaria produced more multiple shoots when were cultured in the liquid proliferation medium, MS medium containing 2.0 mg/L BA and 2.0 mg/L IBA. To maintain the in vitro plantlets of Zingiberaceae species, they were required to subculture every four weeks. After executing proper acclimatization protocol, in vitro plantlets of Alpinia galanga, A. conchigera, Curcuma domestica, C. zedoaria, Kaempferia galanga and Zingiber officinale could be successfully planted in the field with high percentage of survival.

• Korean Journal of Animal Reproduction
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• v.17 no.1
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• pp.69-74
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• 1993

This study was investigated to examine the effect of conditioned medium from bovine oviductal cell(BOEC) in the co-culture system with BOEC on in vitro development of in vitro produced bovine embryos. Oocyte-cumulus complexes were cultured for 24 hrs in TCM-199 supplemented with 10% fetal calf serum, 1$\mu\textrm{g}$/ml FSH and 21U hCG, 1$\mu\textrm{g}$/ml oestradiol-17$\beta$ at 39$^{\circ}C$ under 5% CO2 in air. In vitro fertilization was performed with epididymal sperm and heparin (10$\mu\textrm{g}$/ml, 15min.) or caffeine(2.5mM)-treated spermatozoa. Oocytes were incubated with 1$\times$106 spermatozoa/ml for 18 hrs and then cultured in various culture system for 7 days. The development rates of 16-cell or blastocyst stages were recorded on 4, 7 days, respectively, after incubating. The proportions ofembryonic development into molulae and blastocysts were higher in cumulus cell co-culture(23.4%) and BOEC co-culture(34.3%) than in M199-FCS(6.1%). Similarily, the development rates into molulae and blastocysts were significantly higher in BOEC-conditioned medium than those in M199-FCS. Therefore, it is suggested that BOEC co-culture and BOEC conditioned medium increase significantly the development of in vitro produced bovine embryos in in vitro system.

• Journal of Embryo Transfer
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• v.11 no.3
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• pp.217-223
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• 1996

This study was conducted to investigate the effects of co-culture for the development rate to morula /blastocyst stages of early porcine embryos, derived from oocytes matured and fertilized in vitro, with porcine endometrial cell monolayers(PEM) in the two different media, respectively. The rates of embryos developed to 2-, 4-, 8~16-cell and morula /blastocyst stage were 49.6, 40.5, 28.2 and 15.3% in Ham's F-10 with PEM, and 55.3, 45.9, 32.7, and 17.6% in TCM-HEPES with PEM, respectively. The above development rates to morula /blastocyst stages were significantly higher than those of the embryos cultured in the Ham's F-10 and TGM-HEPES without PEM(P<0.05). The in vitro development rates to the morula /blastocyst stage of 1-cell embryos cultured in Ham's F-10 and TCM-HEPES without PEM were 0~1.2%. Especially, most of embryos were observed to arrest the development beyond 4-cell stages. As shown in the above results, the co-culture of in vitro produced porcine embryos with PEM in the two different media enhanced the development of fertilized eggs to morula /blastocyst stages in vitro. However, we didn't find out any differences for the in vitro development to morula /blastocyst stages between Ham's F-10 and TcM-HEPES media.

• Korean Journal of Animal Reproduction
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• v.18 no.1
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• pp.47-54
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• 1994

The objective of this study was to produce Korean native calves following transfer of in vitro matured, fertilized and cultured embryos into Holstein cows. The ovaries of Korean native cows or heifers were obtained from an abattoir and kept on 25 to 28$^{\circ}C$ and transported to laboratory within 2 hrs. The oocytes were matured in vitro (IVM) for 24 hrs in TCM-199 supplemented with 35$\mu\textrm{g}$/ml FSH, 10$\mu\textrm{g}$/ml LH, 1$\mu\textrm{g}$/ml estradiol-17$\beta$ and granulosa cells at 39$^{\circ}C$ under 5% CO2 in air. They were fertilized in vitro (IVF) by epididymal spermatozoa treated with heparin for 24 hrs., and then the zygotes were co-cultured in vitro (IVC) with bovine oviductal epithelial cells for 7 to 9 days. Late morulae and blastocysts produced in vitro were nonsurgically transferred to recipient cows by unilaterial. Recipients were monitored for estrus and for pegnancy by rectal plapation in 60 days after embryo transfer. One of them was pregnant to term and produced a female weighing 42.5kg at birth.

• Journal of Embryo Transfer
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• v.11 no.2
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• pp.125-135
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• 1996

The present study was carried out to investigate the effects of cryoprotectants, equilibration step, freezing rate, culture condition following in vitro fertilization, and age and development stage of embryo by freezing with conventional slow freezing and vitrification on survival of frozen-thawed Korean native cattle(KNC) blastocysts produced in vitro. The KNC blastocysts produced in vitro were equilibrated in 1.8M ethylene glycol or 1.4M glycerol and cooled from -6$^{\circ}C$ to -35$^{\circ}C$ at -0.3$^{\circ}C$ or -O.6$^{\circ}C$ /minute. When equilibrated in 1.8M ethylene glycol, survival rate of fiozen4hawed blastocysts was sarne in both -0. 3$^{\circ}C$ /min and -0.6$^{\circ}C$ /min cooling rate(71.4%). With the equilibration in 1.4M glycerol, survival rate was higher in -0.3$^{\circ}C$ /min(63.6%) than in -0.6$^{\circ}C$ /min cooling rate(53.8%). For vitrification of the KNC blastocysts produced in vitro, they were equilibrated in 2-step or 3-step exposure to vitrification solution(25% ethylene glycol + 25% glycerol). Survival rate was sirilar in both 2-step(45.0%) and 3-step exposure(47.4%). According to culture condition following in vitro fertilization, higher survival rate was obtained for blastocysts co-cultured with bovine oviductal epithelial cell(BOEC, 77.3%) than for those cultured with epidermal growth factor(EGF, 65.7%) or for those co-cultured with BOEG + EGF (54.8%). According to embryo age and development stage, higher survival rate was obtained for 7-day ernbryos(70.0%) than 8-day(56.8%) or 9-day(20.0%) for blastocyst stage and obtained for 8-day embryos(74.3%) than 7-day(62.5%) or 9-day(42.9%) for exponded blastocyst. In surnmary, higher survival rate of frozen4hawed KNC blastocysts produced in vitro were obtained by using ethylene glycol for cryoprotectant and -0.3$^{\circ}C$ /min for cooling rate. And higher survival rate were obtained with co-culture with BOEC for culture condition following in vitro fertilization and with 7-day blastocyst or 8-day expanded blasto cyst for embryo age and development stage.

• Journal of Animal Reproduction and Biotechnology
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• v.35 no.1
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• pp.86-93
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• 2020

The establishment of porcine embryonic stem cells (ESCs) from porcine somatic cell nuclear transfer (SCNT) blastocysts is influenced by in vitro culture day of porcine reconstructed embryo and feeder cell type. Therefore, the objective of the present study was to determine the optimal in vitro culture period for reconstructed porcine SCNT embryos and mouse embryonic fibroblast (MEF) feeder cell type for enhancing colony formation efficiency from the inner cell mass (ICM) of porcine SCNT blastocysts and their outgrowth. As the results, porcine SCNT blastocysts produced through in vitro culture of the reconstructed embryos for 8 days showed significantly increased efficiency in the formation of colonies, compared to those for 7 days. Moreover, MEF feeder cells derived from outbred ICR mice showed numerically the highest efficiency of colony formation in blastocysts produced through in vitro culture of porcine SCNT embryos for 8 days and porcine ESCs with typical ESC morphology were maintained more successfully over Passage 2 on outbred ICR mice-derived MEF feeder cells than on MEF feeder cells derived from inbred C57BL/6 and hybrid B6CBAF1 mice. Overall, the harmonization of porcine SCNT blastocysts produced through in vitro culture of the reconstructed embryos for 8 days and MEF feeder cells derived from outbred ICR mice will greatly contribute to the successful establishment of ESCs derived from porcine SCNT blastocysts.

• Korean Journal of Animal Reproduction
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• v.18 no.4
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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.