• Journal of The Korean Society of Grassland and Forage Science
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• v.9 no.2
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• pp.62-67
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• 1989

A breeding program in progress at Suweon Livestock Experiment Station, RDA, involves the hybridization of italian ryegrass (Lm) and several leading forage grasses in an effort to combine the nutritive, productivity and palatability qualities of Lm with the adaptive and cold resistance qual; .les of several leading grasses. In order to study the fate of the hybridization between remotely related species, immatured hybrid embryos were cultured on media. The emasculated plants of Lm were Sikem and Tetrone. Reweille, 2n=14, of perennial ryegrass (Lp.), Forager, 2n=42, of tall fescue(Fa), First, 2n=14,of meadow fescue(Fp), Potomac, 2n=28, of orchardgrass(Dg), and Richmond, 2n=42, of timothy(Ph.p) were used as pollinators. Embryos were isolated on 4, 8, 12, 16, and 20 days after pollination and cultured them on modified Murashige and Skoog media. Calluses and plantlets have been obtained after 8 days old embryos crossing between $Lm{\times}Fa$, after 12 days embryos crossing between $Lm{\times}Lp$, Dg, and Ph.p, and after 16 days embryos crossing between $Lm{\times}Fp$. Both callus and shoot formation occurred on 6 % or less of the plated embryos of $Lm{\times}Fp$, Dg, and Ph.p. Embryoderived callus forming shoots have been obtained from 4 days old embryo crossing between $Lm{\times}Dg$ which has not successfully been done anywhere as far as we know. It means that hybrid plants of species without crossability can be obtained through the use of immatured embryo culture. Some of plated embryos developed directly shoots from embryos and the others shoot-forming callus. Cross between related species showed a high frequency of directly shoot formation from plated embryos and cross between remotely related species a high frequency of callus formation.

• Korean Journal of Animal Reproduction
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• v.10 no.1
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• pp.27-35
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• 1986

As a preliminary experiment to establish the process on the sexing of mouse embryos by chromosomal analysis, present studies were carried out with inbred (ICR, C57BL) and F1 hybrid [(ICR${\times}$C57BL) = F1 ${\times}$ ICR] mice to investigate the blastomere numbers and mitotic indices (M.I.) to the developmental stage of embryos recovered, the optimum periods of anti-mitotic agent administration, the successful rates of sexing and sex-ratio. The results obtained were summarized as follows: 1. The blastomere numbers (mean${\pm}$S.E.) of the morula and blastocyst were 18${\pm}$0.4 and 54${\pm}$0.7, respectively. 2. Whereas the M.I. of F1 hybrid (16${\pm}$0.2%) was higher than that fo inbred ICR (15${\pm}$0.1%) and C57BL (12${\pm}$0.6%) in the different strains, the morula (7${\pm}$0.6%) was higher than that of blastocyst (6${\pm}$0.4%) in the case of embryo stages. 3. Following to anti-mitotic agents treated, the M.I. of embryos cultured with Colcemid (17${\pm}$1.1%) was superior to that fo embryos cultured with Velban (12${\pm}$0.9%) and the Colcemid injection (7${\pm}$0.4%). 4. The successful rate of sexing in the blastocyst (38.7%; 124/320) was superior to the morula (35.9%; 52/145), and the F1 hybrid (48.1%) was higher than that of inbred ICR (42.4%) and C57 BL (28.2%). 5. In the successful rate of sexing to the methods of administration, the embryos cultured with Colcemid (46.0%) was superior to that of embryos cultured with Velban (39.0%) and the Colcemid injection (38.8%). 6. Of 98 embryos sexed after culture with Colcemid, 89(90.8%) were observed between 2 and 4 hrs. In the case of Velban treatment, 83.1% (74/89) was observed between 2$\frac{1}{2}$ and 4$\frac{1}{2}$ hrs. 7. Out of 761 prepared embryos it was possible to sex 311; 157 were male and 154 were female, i.e.a sex-ratio of 50% a, pp.oximately.

• Clinical and Experimental Reproductive Medicine
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• v.19 no.2
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• pp.117-123
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• 1992

Development in vitro of 2-cell mouse embryos was examined after appropriate exposure to oviductal milieu to demonstrate biological activity present in the oviducts. ICR and ($C57Bl/6{\times}Balb/c$) $F_1$ hybrid mice were superovulated and mated for the recovery of early embryos. Embryos were recoverd at every 2h intervals from 32h post-hCG(hph) to 56 hph. The proportions of developmental stages were determined in the recovered embryos. Development in vitro of 2-cell embryos was more rapid in $F_1$ hybrid than in ICR, showing high proportions of 4-cell embryo and blastocyst at 120 hph. 100% of blastocyst development was obtained at 38hph in $F_1$ hybrid and at 50 hph in ICR when 2-cell embryos were cultured upto 120hph in vitro. Moreover, in vitro culture of oviducts containing 2-cell embryos in ICR mice for 12h from 34hph to 46hph increased developmental capacity of ICR mouse embryo in vitro. The results indicate that oviductal environment contains substances having mitogenic activity and overcoming early cell block in vitro. The mitogenic activity is effective in vitro as well as in vivo.

• The Korean Journal of Zoology
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• v.34 no.3
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• pp.389-393
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• 1991

To obtain monozygotic multiplets from 8-cell mouse embryos, we artificially constructed chimeric embryos by introducing one blastomere (donor) of 8-cell embryos of Fl hybrid (C57BL/6 X CBA) mice into 4-cell ICR mouse embryos (carrier) of which one blastomere had been previously removed with a micromanipulator. After 42 h of culture, the developmental frequency of chimeric embryos to normal morula and blastocyst was 95% (310/328). When chimeric embryos at morula or blastocvst stage were transferred to pseudopregnant mice,39%, (70/180) of them were born. Most of the offspring (56/70) were the carrier type in coat color, whereas only three of them were the donor type, of which ho were assumed to be derived from single 8-cell donor embryo. Because the two donor type mice Ivere the same sex and produced only the donor type offspring from a testcross, they are probably monozvgotic multiplets of 8-cell mouse embryos. However, since their internal chimerism was not able to be examined, it remains to be determined if their genetic constitutions are identical.

• Journal of Plant Biotechnology
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• v.33 no.2
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• pp.99-104
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• 2006

This study was performed to investigate regeneration of plants differentiated from hybrid embryos between L. longifilorum Georgia and L. elegans Kakutanohikari. In addition, proliferation of callus and process of differentiation were investigated by histological observation. The germination of hybrid embryos was observed in 86 individuals from 48 slice cultures. Plant regeneration was effective on a medium supplemented with 1 mg/L HPh, and only callus proliferation was the highest in combination of 0.1 mg/L HPh and 1 mg/L BA. Also, plant regeneration was the most effective on a medium supplemented with 50 mg/L pyridoxine. We concluded that somatic embryos were formed from procambium of callus and proliferation of embryonic or proembryonic cells were stimulated with NAA from procambial cells.

• The Korean Journal of Zoology
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• v.32 no.4
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• pp.404-411
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• 1989

In this study, we attempted to determine the precise patterns of protein synthesis during the second cleavage in mouse. F1 hybrid 2-cell embryos showing a highly synchronized cell cycle and outbreed ICR strain 2-cell embryos were used. The patterns of protein synthesis during the second cleavage showed the sequential changes in the F1 hybrid and ICR strain 2-cell embryos. Moreover, we examined the effects of mitotic and transcriptional inhibitors such as colcemid and a-amanitin on the protein synthesis in the late 2-cell embryos of ICR strain. Treatment of colcemid (0.1mg/ml) blocked the second cleavage, but did not affect on the change of protein synthesis. However, treatment of a-amanitin induced the synthesis of two set of polypeptides without affecting on synthesis of other proteins and cleavage. It thus seems that the appearance of a-amanitin-sensitive proteins may be not involved in the second cleavage. Therefore, these results indicate that the second cell cycle in mouse embryos appears to be regulated at post transcriptional level, presumably independent on the expression of embryonic genome. 본 연구는 생쥐 배아의 2세포기 분열과정중 단백질 합성양상과 단백질합성에 미치는 colcemid와 $\alpha$-amanitin의 영향을 조사하였다 이를 위하여 체내 수정된 ICR strain의 2세포기 배아와 매우 일치된 초기배아 분열양상을 보여주는 체외 수정된 F1 (C57BL x CBA) hybrid 2세포기 배아를 사용하였다. 두 종류의 2세포기 배아에서 단백질 합성은 분열단계에 따라서 매우 일치된 변화를 보여 주었다. 또한 유사분열 억제제인 colcemid (0.1mg/ml)의 처리는 2세포기 배아분열을 억제하였으나, 단백질 합성에는 아무런 변화를 주지 못하였다. 그리고 후기 2세포기 배아에 전사 억제제인 a-amanitin (100mg/ml)을 처리하였을 때 세포분열이나 다른 단백질의 합성에는 아무런 영향이 없이 단지 두개의 단백질의 합성만을 유도하였다. 이는 아마도 a-amanitin의 stress효과에 기인하는 것으로 추측된다. 따라서 생쥐 2세포기 배아의 분열과정은 배아게놈의 유전자 발현과는 무관하게 이미 합성되어 존재하는 mRNA에 의하여 조절되는 것으로 사료된다.

• Journal of Ginseng Research
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• v.43 no.1
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• pp.38-48
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• 2019

Background: Interspecific ginseng hybrid, Panax ginseng ${\times}$ Panax quenquifolius (Pgq) has vigorous growth and produces larger roots than its parents. However, F1 progenies are complete male sterile. Plant tissue culture technology can circumvent the issue and propagate the hybrid. Methods: Murashige and Skoog (MS) medium with different concentrations (0, 2, 4, and 6 mg/L) of 2,4-dichlorophenoxyacetic acid (2,4-D) was used for callus induction and somatic embryogenesis (SE). The embryos, after culturing on $GA_3$ supplemented medium, were transferred to hormone free 1/2 Schenk and Hildebrandt (SH) medium. The developed taproots with dormant buds were treated with $GA_3$ to break the bud dormancy, and transferred to soil. Hybrid Pgq plants were verified by random amplified polymorphic DNA (RAPD) and inter simple sequence repeat (ISSR) analyses and by LC-IT-TOF-MS. Results: We conducted a comparative study of somatic embryogenesis (SE) in Pgq and its parents, and attempted to establish the soil transfer of in vitro propagated Pgq tap roots. The Pgq explants showed higher rate of embryogenesis (~56% at 2 mg/L 2,4-D concentration) as well as higher number of embryos per explants (~7 at the same 2,4-D concentration) compared to its either parents. The germinated embryos, after culturing on $GA_3$ supplemented medium, were transferred to hormone free 1/2 SH medium to support the continued growth and kept until nutrient depletion induced senescence (NuDIS) of leaf defoliation occurred (4 months). By that time, thickened tap roots with well-developed lateral roots and dormant buds were obtained. All Pgq tap roots pretreated with 20 mg/L $GA_3$ for at least a week produced new shoots after soil transfer. We selected the discriminatory RAPD and ISSR markers to find the interspecific ginseng hybrid among its parents. The $F_1$ hybrid (Pgq) contained species specific 2 ginsenosides (ginsenoside Rf in P. ginseng and pseudoginsenosides $F_{11}$ in P. quinquefolius), and higher amount of other ginsenosides than its parents. Conclusion: Micropropagation of interspecific hybrid ginseng can give an opportunity for continuous production of plants.

• Clinical and Experimental Reproductive Medicine
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• v.28 no.1
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• pp.55-63
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• 2001

Objectives: This study was carried out to evaluate the effects of embryonic stage, cryoprotectant, and freezing-thawing method on the rates of survival and development of the cryopreserved mouse early embryo and finally to establish the cryopreservation method of surplus embryos obtained during assisted reproductive technology (ART). Materials and Methods: Two to eight cell embryos were obtained from oviducts of mated $F_1$ hybrid female mice superovulated by pregnant mare's serum gonadotropin (PMSG) and human chorionic gonadotropin (hCG). Two-step 1,2-propanediol (PROH), dimethylsulfoxide (DMSO) and 4-step PROH DMSO were used as cryoprotectant and dehydration and rehydration method of embryos, and slow-cooling or rapid-cooling method was used as frozen program. The survival rates of embryos were measured after thawing and rehydration, and the developmental rates of embryos were compared and observed during culturing embryos for 24, 48, 72, 96 hrs. Results: As for the survival and development rates of embryos according to embryonic stage, the survival rate of 2 cell stage in PROH and DMSO was significantly higher than 4-8 cell (64.5% versus 62.1 %,79.7% versus 73.2%) (p<0.01, p<0.01), but the development rates of 4-8 cell embryos in PROH and DMSO were significantly higher than 2 cell embryos for whole culture period (p<0.01) and the development rates of 4-8 cell embryos in PROH were significantly higher than 2 cell embryos in DMSO (p<0.01). As for the survival and development rates of embryos according to cryoprotectant, the survival rate of 2 cell embryo in DMSO was significantly higher than that in PROH (74.4% versus 64.5%) (p<0.01), whereas the development rate of embryos was not differ till 24 hrs. The developmen1 rate from morular to hatching blastocyst, however, was significantly higher in PROH than in DMSO during 48 hr (p<0.01). The survival rate of 4-8 cell embryo was 62.1% in PROH and 73.2% in DMSO. The development rates of embryo in PROH were significantly higher for whole culture periods (p<0.01, 0.05). In respect to the effect of freezing and thawing program on the survival and development rates of embryos, method of slow cooling and rapid thawing was more effective than that of rapid cooling and rapid thawing. Conclusions: The survival rate of embryo in 2 cell stage was higher than in 4-8 cell stage, and PROH appears more effective cryoprotectant than DMSO because PROH showed better development rates of embryos in 2 and 4-8 cell stage. Moreover, slow cooling and rapid thawing method was considered as the best cryopreservation program.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.52 no.3
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• pp.298-301
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• 2019

We investigated the development of allotriploid embryos derived from a cross between female starry flounder Platichthys stellatus and male stone flounder Kareius bicoloratus. The second cleavage, mid-blastula, gastrula, and Kupffer's vesicle appearance stages, and hatching of embryos began 3.7, 25.6, 45.7, 87.7, and 213.2 h after cold shock at $6^{\circ}C$, respectively. The hatching and development time of triploid interspecific hybrid eggs was approximately the same as those of diploid starry flounder eggs at the same incubation temperature.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.52 no.3
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• pp.274-282
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• 2019

Female Siberian sturgeon Acipenser baerii were crossed with male Russian sturgeon A. gueldenstaedtii and the development, ontogenetic behavior, and phototactic characteristics of the resultant hybrid prolarvae were examined. The fertilization rate of the hybrid group was similar to that of Siberian sturgeon (the maternal species), and the overall developmental characteristics of hybrid embryos were not significantly different from those of either parental species. The time window from first hatch to the completion of hatching was wider in Russian sturgeon than in Siberian and hybrid sturgeon groups. The prolarval viability of the hybrid group was similar to that of the maternal species, and significantly higher than that of the paternal species. Hybrid prolarvae displayed ontogenetic behavioral patterns that were quite similar to those of Siberian sturgeon. In an illumination preference test, hybrid prolarvae displayed significant positive phototaxis, similar to the maternal species, and clearly distinct from the negative phototactic character of the paternal species. Taken together, our data indicate that the prolarval traits of hybrid sturgeon from this cross more closely resemble those of the maternal than the paternal species.