• Korean Journal of Plant Tissue Culture
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• v.22 no.5
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• pp.291-298
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• 1995

This study describes plant regeneration from leaf explant of Bupleurum falcatum through somatic embryogenesis, and the effect of 2,4-dichlorophenoxyacetic acid on somatic embryo abnormalities. The relationship between the cotyledon number of somatic embryo and its germinability is also described. Embryogenic calli were selected from calli formed on explants cultured on MS solid basal medium supplemented with 1 mg/L 2,4-D. Cotyledonary abnormalities were observed in somatic embryos which were developed from calli cultured on MS medium with 1 mg/L 2,4-D for 6-week and then subcultured on 2,4-D free MS medium for 3 weeks. The frequency of abnormalities was as follows: 7% of somatic embryos had one cotyledon, 65% of them had two cotyledons, 25% three cotyledons, 5% four cotyledons, 2% five cotyledons, and 3% trumpet-like cotyledons. The two cotyledon somatic embryos were germinated at a frequency of 80%. However the germination frequency of one cotyledon embryo and multicotyledonary embryo was lower than that of the two cotyledon somatic embryo. All of trumper-like somatic embryos did not germinate. Histological observations of multicotyledon embryo showed circular procambium in the root but pocambial strands in the cotyledonary node or upper hypocotyl. The number of the strands was equal to the cotyledon number.

• Korean Journal of Medicinal Crop Science
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• v.3 no.2
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• pp.100-106
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• 1995

This study was conducted to find the factors affecting somatic embryogenesis in suspension culture of Rehmania glutinosa and investigate the possibility of artificial seed production by encapsulation of somatic embryos. Linsmeier-Skoog medium was appeared as proper for somatic embryogenesis. Sucrose with $3{\sim}5%$ as carbon sources was good for somatic embryogenesis, and both ammonium and nitrate nitrogen were necesary for normal somatic embryo production. BA with NAA or kinetin with NAA were better than the use of cytokinin alone for both somatic embryogenesis and numbers of somatic embryos. $AgNO_3$ as protectant for vitrification of seedlings in vitro culture had no harmful effect on somatic embryos. Sphericity of encapsulated seeds was good at 3% gel of sodium alginate but germination was better at 2.5% sodium alginate level. Artificial seeds were germinated and developed normal shoots and roots under in vitro condition.

• Korean Journal of Animal Reproduction
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• v.18 no.1
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• pp.7-13
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• 1994

To verify in vivo viability of IVF-derived bovine embryos, morula and blastocysts that developed from in vitro matured and fertilized ova were transferred to the uteri of recipient cows and normal calves were produced. To produce IVF-derived bovine morula or blastocysts, ova matured and fertilized in vitro were cultured in culture medium for 7~8 days at 39$^{\circ}C$ under the humicified atmosphere of 5% CO2. Two different culture systems, a co-culture system with TCM-199 and bovine epithelial cells (BOEC) and CR1aa without somatic cell support, were compared. Cleavage rates to 2~8 cell stage and developmental rates of IVF-derived bovine embryos to blastocyst stage were not different between co-culture system (51.3 and 14.0%) and CR1aa medium (60.4 and 22.1%), respectively. Embryos were classified into three grades by embryo quality and then one or two embryos in higher quality(A and B grades) were transferred to the uterus of recipients. In this study Korean Native calf was first born after transfer of IVF-derived embryos. Total four live calves were normally developed to term from IVF-derived bovine blastocysts and one female fetus was still-born approximatedly 8 months of gestation, but there was no pregnancy after transfer of morula. Therefore, normal calves could be produced after transfer of IVF-derived bovine embryos cultured in CR1aa medium without somatic cell support. In addition, our results suggest that in transfer of IVF-derived bovine embryos blastocyst stage is better than morula.

• Korean Journal of Plant Tissue Culture
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• v.26 no.4
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• pp.289-291
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• 1999

Hypocotyl explants from 7 days old seedlings of one $F_1$ hybrid cultivar and two pure lines of cucumber formed embryogenic calli at frequencies of up to 8% when cultured on Murashige and Skoog medium (MS) supplemented with 1 mg/L 2,4-D for 3 weeks. Embryogenic calli gave rise to somatic embryos. When slices of somatic embryos were cultured on the same medium for 4 weeks, they formed embryogenic calli. Embryogenic cell suspension cultures were established with embryogenic calli in MS liquid medium with 1 mg/L 2,4-D. Embryogenic potential of cell suspension cultures was maintained by subculturing every seven days. When the level of 2,4-D in the medium was lowered to 0.2 mg/L by diluting with liquid MS basal medium, embryogenic cell suspension cultures underwent development into numerous somatic embryos. When plated onto MS basal medium, over 95% of somatic embryos developed into plantlets. Plantlets were transplanted to potting soil and grown to maturity.

• Journal of Animal Reproduction and Biotechnology
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• v.34 no.3
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• pp.232-239
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• 2019

In general, cloned pigs have been produced using the somatic cell nuclear transfer (SCNT) technique with various types of somatic cells; however, the SCNT technique has disadvantages not only in its low efficiency but also in the development of abnormal clones. This study aimed to compare early embryonic development and quality of SCNT embryos with those of induced pluripotent stem cells (iPSCs) NT embryos (iPSC-NTs). Ear fibroblast cells were used as donor cells and iPSCs were generated from these cells by lentiviral transduction with human six factors (Oct4, Sox2, c-Myc, Nanog, Klf4 and Lin28). Blastocyst formation rate in iPSC-NT (23/258, 8.9%) was significantly lower than that in SCNT (46/175, 26.3%; p < 0.05). Total cell number in blastocysts was similar between two groups, but blastocysts in iPSC-NT had a lower number of apoptotic cells than in SCNT (2.0 ± 0.6 vs. 9.8 ± 2.9, p < 0.05). Quantitative PCR data showed that apoptosis-related genes (bax, caspase-3, and caspase-9) were highly expressed in SCNT than iPSC-NT (p < 0.05). Although an early development rate was low in iPSC-NT, the quality of cloned embryos from porcine iPSC was higher than that of embryos from somatic cells. Therefore, porcine iPSCs could be used as a preferable cell source to create a clone or transgenic animals by using the NT technique.

• Journal of The Korean Society of Grassland and Forage Science
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• v.19 no.4
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• pp.297-302
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• 1999

Cotyledonary abnormalities were observed in secondary somatic embryos which were developed from calli cultured on MS medium with various concentrations of 2,4-D. In MS medium containing hormone-free or $0.1mg/{\ell}$ 2,4-D, the frequency of normal embryo formation with two cotyledons were above 57%. According to concentration of 2,4-D increment the frequency of normal embryos were decreased. In MS medium containing $4mg/{\ell}$ 2,4-D, the frequency of normal embryo formation was just 10%. The rate of germination was as follows; 37.7% of somatic embryos had one cotyledon, 85% two cotyledons, 38% three cotyledons, 35% four cotyledons, 25% five cotyledons and 29% trumpet-like cotyledons. About 80% of the embryos with two cotyledons were converted into normal plants, but one, three or four cotyledons were only 6.8~10%. The five or trumpet-like embryos were not developed into normal plants.

• Journal of Veterinary Clinics
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• v.16 no.1
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• pp.163-176
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• 1999

The nuclear transplantation technique is known as the most potential and efficient method for producing large numbers of genetically identical animals from a single embryo and somatic cells. After Dolly was introduced in 1997, many scientists were amazed. A possibility came to a reality that live offspring could be produced with differentiated somatic cells from an adult animal. On the other side, many in the press and the sensationalists focused on the socially, ethically and scientifically unacceptable sides of the technology. In this article, the history, current status and prospects of the technological development of nuclear transplantation in mammals and its application to the production of cloned animals are described. For the efficient and successful production of cloned embryos by nuclear transplantation, the right selection, preactivation and micromanipulation of oocytes as capacious recipient cytoplasm, the adequate and benefitial preparation of multiple totipotent embryonic and somatic cells as donor nuclei, fusion of them and in vitro production of cloned embryos are very critical. Recently the overall efficiency of production of cloned embryos and offspring in livestock has been much improved. Cloning will also be a more efficient, faster and useful way of creating transgenic fetuses for gene therapies, gene pharming, organs for xenotransplantation by preselection and mass production of transgenic embryos and consequently improving the production efficiency in transgenic animals. Further technical development of nuclear transplantation will enable large-scale production of cloned livestock and in near future the commercial cloning of animals will become a reality.

• Proceedings of the Korean Society of Developmental Biology Conference
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• 1998.07a
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• pp.43-44
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• 1998

Effect of somatic cell coculture on hatching of mouse blastocyst was examined. Mid-blastocysts were cocultured with granulosa cell primary culture or Sertoli cell line ($TM_{4}$) derived from mouse testis for 48 hr. Blastocysts cultured in medium (10% FBS) started to hatch more faster than cocultured embryos during 12 hr of coculture. After then blastocysts cocultured with somatic cell hatched faster than control. Degeneration of embryos was also greately reduced by coculture. This result suggested the potentiation of hatching as well as embryonic viability by coculture with somatic cell and Sertoli cell line can be used for embryo coculture.

• Korean Journal of Plant Tissue Culture
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• v.27 no.2
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• pp.149-154
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• 2000

Embryogenic suspension cultures were initiated using embryogenic callus from immature inflorescence explants (Aralia cordata Thunb.) cultured on solid MS medium containing 1 mg/L 2,4-D for 8 weeks and then the embryogenic callus was proliferated in liquid MS medium containing 1 mg/L 2,4-D. After sieving the suspensions (pore size 270$\mu$m), embryogenic cells were cultured in liquid MS medium with cytokinins (kinetin, BA, zeatin) for two weeks. When the embryogenic cells were transferred to liquid MS basal medium, primary somatic embryos were developed after 5 weeks of culture. Secondary embryos were developed directly from the primary torpedo and cotyledonary embryos cultured in solid MS basal medium. Frequency of secondary embryogenesis was higher on medium containing 2 mg/L kinetin than the other cytokinins. Plant regeneration was highly recorded by placing secondary cotyledonary embryos induced from primary cotyledonary embryos in MS medium containing 2 mg/L kinetin or 2 mg/L zeatin (25.4% and 28.6%, respectively). The plant regeneration from secordary embryos was prohibited by tertiary embryogenesis.

• Journal of Embryo Transfer
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• v.23 no.1
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• pp.31-36
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• 2008

We attempted to control the maturation promoting factors (MPF) activity and nuclear remodeling of somatic cell nuclear transfer (NT) bovine embryos. Bovine ear skin fibroblasts were fused to enucleated oocytes treated with either 5 mM caffeine for 2.5 h or 0.5 mM vanadate for 0.5 h and activated. The nuclear remodeling type of the reconstituted embryos was evaluated 1.5 h after activation. MPF activity was assessed in enucleated and chemical treated oocytes before the injection of a donor cell. Effect of chemicals on the embryonic development was evaluated with parthenogenetic embryos. MPF activity increased significantly by caffeine treatment, but decreased by vanadate treatment (p<0.05). Caffeine or vanadate had no deleterious effect on the parthenogenetic embryo development. In caffeine treated group, premature chromosome condensation (PCC) was occurred in 72.2% of NT embryos (p<0.05). In contrast, vanadate induced the formation of a pronucleus-like structure (PN) in a high frequency (68.9%, p<0.05) without PCC (NPCC). Blastocyst development of NT embryos increased by treating with caffeine (30.3%), whereas decreased by treating with vanadate (11.4%) compared to control (22.1%, p<0.05). The results indicate that caffeine or vanadate can control of MPF activity and remodeling type of NT embryos, resulting in the increased or decreased in vitro development.