• Korean Journal of Veterinary Research
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• v.30 no.3
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• pp.317-331
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• 1990

Chromosomes of gonadal tissues from Fasciola spp, Eurytrema pancreaticum and Calicophoron calicophorum occurred Korean cattle were egamined using modified air-drying method. To compare their phenotype with three different genotypes among Fasciola spp, the adult and egg si2e were measured since they have been known as important taxonomical characters. The results obtained were as followed; Cattle liver fluke, Fasciola spp were classified into three types based on their chromosomal complements such as individual with 2o chromosome(diploid), 30 chromosome(triploid) and 20/30 mosaic constitution(mixoploid). The propotions of appearance of three types were 40.00%, 54.29% and 5.71%, respectively. The frequency of three types in type I which was regarded as F gigantica were 58.82% for diploid, 35.29% for triploid and 5.88% for mixoploid, but in type II which was regarded as F hepatica were 72.2% for triploid, 22.22% for diploid and 5.56% for mixoploid. Egg length of triploid forms was significantly larger than that of diploid forms and egg size of mixoploid forms was similiar to that of triploid forms. Worm size of triploid forms was larger than that of diploid forms and was more similar to that of mixoploid forms, but the statistical data were not significant. Diploid chromosome consisted of one pair of metacentric chromosome(m), four pairs of submetacentric chromosomes(sm), five pairs of subtelocentric chromosomes(st), while triploid chromosome consisted of one pair of metacentric chromosome, seven pairs of submetacen.tric chromosomes, one pair of subtelocentric chromosome and telocentric chromosome(t), respectively. In mixoploid chromosome, constitution of the chromosomes of diploid or triploid cell was consistent with that of diploid or triploid. Chromosomes of gonadal tissues from pancreatic fluke, Eurytrema pancreaticum consisted of 13 pairs of homologs(2n=26, n=13). The mitotic and meiotic divisions were observed frequently. In the mitotic metaphase, Karyotype consisted of five pairs of metacentric chromosomes, four pairs of submetacentric chromosomes, three pairs of subtelocentric chromosomes and one pair of telocentric chromosome. Chromosomes of gonadal tissues from stomach fluke, Calicvphoron calicophorum consisted of 9 pairs of homologs(2n=18, n=9). The meiotic divisions was frequently observed, but mitotic divisions was rare. In the mitotic metaphase, Karyotype consisted of two pairs of metacentric chromosomes, three pairs of submetacentric chromosomes and four pairs of subtelocentric chromosomes. Karyotype of Calicophoron calicophorum differed from that of Japanese C calicophorum which was similar to that of Paramphistomum cervi of Korean cattle. Though that of Calicophoron calicophorum of Korean cattle was similar to that of Paramphistomum explanatum of Korean cattle, that have been recognized to be a different species of fluke.

• Parasites, Hosts and Diseases
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• v.25 no.1
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• pp.37-44
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• 1987

As a series of systematic classification for Korean common liver fluke, Fasciola sp., karyotype was investigated by means of the modified air-drying technique and of the regular Giemsa staining. Also, C-staining method was applied for detailed karyological analysis from the germ cells of the fluke. The following is a brief summary of the leading facts gained through the experiment. 1. Korean Pasciola sp. was classified into three types based on their chromosomal complements; individuals with 20 or 30 chromosomes and with a 20/30 mosaic constitution. Worms having 30 chromosomes represent a triploid form with 3 sets of 10 basic chromosomes, while those with 20 chromosomes were diploid and mosaic individuals were 2n/3n mixoploid. 2. The frequency of the individual type calculated is as follows; 67.45% of 212 flukes examined was of diploid, 10.85%, triploid, and the rest, 21.7%, mixoploid, respectively. In many cases, two or three types were found in the peculiar bovine host while single type inhabitant was about 20% out of 52 cases. 3. The twenty chromosomes consisted of 1 pair of large metacentrics, 4 pairs of medium-sized subtelocentrics, and 5 pairs of small submetacentrics, while constitution of the thirty chromosomes was nearly interpreted as a triploid form with 3 sets of 10 basic chromosomes. The high centromeric indexes of both types are the first Pairs among all the examined, and 37.93% was of diploid and 47.93%, triploid, respectively. 4. In mixoploid individuals, constitution of the chromosomes of diploid or triploid cells was the same as that of diploid or triploid individuals. 5. All the chromosomes of the germ cells in both types showed C-band around the centromeric region and especially the chromosomes no's 3,7, and 8 showed a remarkable C-band distinguished from other chromosomes. 6. The variance for the sizes of the worms and the eggs were not parallel with three different genotypes in Korean common liver fluke.

• Journal of Plant Biology
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• v.39 no.4
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• pp.231-235
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• 1996

The calli obtained from the bulbs of A. victorialis var. platyphyllum on MS based medium containing 2 mg/L 2, 4-D and 1mg/L BA. Plants from calli were regenerated on MS basal media supplemented with combinations of NAA and four kinds of cytokinin, BA, zeatin, kinetin and 2iP, and with only each cytokinin. The good response of shoot regeneration was observed in the media with combinations of NAA and BA or zeatin, and with only BA or zeatin. Shoot regenerating response in the medium with combinations of NAA and BA or zeatin, and with only BA or zeatin. Shoot regerating response in the medium with combinations of NAA and BA or zeatin was about two times higher than that in the mediuim with only BA or zeatin. From the karyotypic analysis of the regenerated plants, abnormal diploids, aneuploids and mixoploid with structural aberrations were investigated. The somaclonal variants (AV 16-04, AV 13-03) were shown the considerable differences from normal diploid regenerant (AV 18-03) in the external morphology.

• Proceedings of the Korean Society of Developmental Biology Conference
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• 2003.10a
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• pp.92-92
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• 2003

Accurate analysis of nuclear status is needed when biopsied-blastomeres are used for embryo sexing. In this study, the nuclear status of blastomeres derived from 8- to 16-cell stage IVF bovine embryos was analyzed to evaluate the representative of single blastomere for embryo sexing. When 55 embryos were analyzed by PCR following biopsy, the coincident rate of sex determination between biopsied-single blastomere and matched blastocyst by PCR was 80 %. Karyotyping of biastomeres in 8- 16-cell stage bovine embryos was conducted to assess chromosome status of IVF embryos. To establish karyotyping of blastomeres, concentrations of vinblastine sulfate and duration of exposure time for metaphase plate induction with 8- to 16-cell stage bovine embryos were tested. The most effective condition for induction of metaphase plate (>45%) was 1.0 ug/ml vinblastine sulfate treatment for 15 h. In 22 embryos under the condition, only 8 embryos out of ten that had a normal diploid chromosome complement showed a sex-chromosomal composition of XX or XY (36.4%) and 2 diploid embryos showed mosaicism of the opposite sex of XX and XY in blastomeres of embryo (9.1%). One haploid embryo contained only one X-chromosome (4.5%). Four out of the other 11 embryos having a mixoploid chromosomal complement contained haploid blastomere with wrong sex chromosome (18.2%). These results suggested that morphologically normal bovine embryos derived from IVF had considerable proportion of mixoploid and sex-chromosomal mosaicism which could be the cause of discrepancies of the sex between biopsied-single blastomere and matched blastocyst by PCR analysis.

• Reproductive and Developmental Biology
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• v.31 no.4
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• pp.273-278
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• 2007

This study was conducted to examine the protein kinase inhibitors, 6-dimethylaminopurine (DMAP) and cycloheximide (CHXM) on the development and chromosome constitution of porcine parthenogenetic embryos. In vitro matured oocytes were activated by electric stimuli (ES) or a combination of ES with culture in 2 mM DMAP or $10{\mu}g/ml$ CHXM for 4 hr. Activated oocytes were cultured in PZM-3 for 6 days. Some 1-cell embryos and blastocysts were fixed by air dry method to analyze the chromosome constitutions and/or total cell number. Blastocyst development of DMAP-treated group (26.7%) was significantly higher (p<0.05) than those of CHXM-treated and ES control groups. Ploidy in 1-cell stage embryos was not different among groups (77.3 to 81.0%), however, proportion of diploid chromosome constitutions was high in DMAP-treated group (61.9%, p<0.05). In the blastocyst stage, proportion of diploid chromosome plates was significantly high in DMAP-treated group (64.2%, p<0.05), and proportion of abnormal chromosome plates was higher in CHXM-treated group (36.6%, p<0.05) than DMAP-treated group (28.3%,). Proportion of embryos with abnormal chromosome constitutions was slightly increased by DMAP (40.0%) and CHXM (42.1%) treatment due to the increasing of mixoploid (47.4 and 52.0%). The present study shows that the DMAP treatment increase the development of porcine parthenotes. However, parthenogenetic activation by ES or combined treatment with ES and DMAP or CHXM detrimentally affects the chromosome constitutions of porcine parthenotes during early embryonic development, leads to increased abnormal ploidy in the blastocyst stage.

• Asian-Australasian Journal of Animal Sciences
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• v.20 no.5
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• pp.718-724
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• 2007

Considerable attention has been focused on the cloning of mammalian embryos, as a consequence of poor development, in order to enhance the application of genetic engineering. Experiments were conducted to compare the developmental competence of parthenotes and reconstructed (NT) rabbit eggs with fetal fibroblasts (FFs) following various activation regimens. Oocytes and NT eggs were exposed to: electric stimulation (EST, Group 1) and EST followed by 6-dimethylaminopurine (DMAP, Group 2), cycloheximide (CHX, Group 3) or DMAP/CHX (Group 4). Pronuclear (PN) status, cleavage, blastocyst development and the ploidy were assessed. In parthenote groups 1, 2, 3 and 4, the PN formation differed significantly. And, the cleavage and blastocyst rates were 41.7 and 5%, 75.6 and 53.7%, 68 and 36%, 82.1 and 52.6%, respectively, among treatments. Polyploidy was observed in 17.2% of EST plus DMAP and 44.9% of EST plus DMAP/CHX groups. In SCNT groups (Group 1, 2, 3 and 4), the cleavage and blastocyst rates were 28.6 and 7.1%, 58.3 and 29.2%, 56.8 and 24.1%, 64.5 and 27.8%, respectively. The chromosomal composition differed significantly (p<0.05) among treatments. In Group 2 and 3, 53.8% and 81.8% of embryos revealed diploid chromosomal sets, respectively. However, in Group 4, 53.3% of embryos showed abnormal ploidy (mixoploid). Although DMAP or combination with DMAP/CHX resulted in higher in vitro development of rabbit SCNT embryos, higher incidence of chromosomal abnormality may induce problems related to fetal loss of at late stage of development.

• Proceedings of the Korean Society of Embryo Transfer Conference
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• 2002.11a
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• pp.77-77
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• 2002

본 연구는 체세포 및 체세포 핵이식란의 염색체 이상에 체세포의 계대의 영향성을 조사하고자 하였다. 한우 성축의 귀 조직으로 얻어진 세포를 공여세포를 체외성숙 후 제핵된 난자에 핵이식을 실시하였으며, 1.9kv/cm, 20$\mu\textrm{s}$/2times의 전기자극으로 융합후 5$\mu\textrm{g}$/ml의 ionomycin에서 4min, 1.9mM 6-DMAP에서 4h동안 배양함으로써 활성화를 유도하였다. 핵이식란은 CRlaa에서 4일간 배양 후 8 -cell단계에서 중기상의 유도를 위하여 상기 배양액 1ml당 0.05$\mu\textrm{g}$ colcemid에서 6-8시간 더 배양하였다. 이후, 6% Fetal bovine serum이 함유된 1% sodium citrate용액에 20분간 저장처리 후, methanol 5 : aceticacid 1 : distilled water 4로 1차, methanol 3: aceticacid 1 로 조성된 2차, methanol 4 : acetic acid 3 : distilled water 1의 3차고정액으로 1분간 재 침지시켰다. 고정 처리가 완료된 slide는 4% Giemsa용액으로 염색한 후 광학현미경 하에서 핵형 양상을 검경하였다. 체세포의 5계대에서는 684개의 spreads를 검경한 결과 염색체 수는 72%가 정상으로 60개이었고, 24%가 60개 이하였으며 4%가 60개 이상을 보였다. 10계대도 5계대와 비슷하여 71%가 정상, 26%가 60개 이하, 3%가 60개 이상이었고, 15계대에서는 55%가 정상이었고, 30%가 60개이하, 15%가 60개 이상을 보였다. 10계대 까지는 mixoploid의 비율의 변화가 없었으나 15계대에서 현저하게 늘어남을 볼 수 있었다. 또한 체외수정란과 핵이식란의 비교에서는 체외수정란은 250개의 spreads를 검경한 결과 염색체 수는 95.6%가 정상으로 60개이었고, 2.0%가 60개 이하, 2.4%가 60개 이상이었으나, 핵이식란은 204개를 검경하여 88%가 정상이었고, 4.9%가 60개이하, 7.1%가 60개 이상을 보임으로써, 핵이식란이 체외수정란에 비하여 염색체 이상의 비율이 높았다. 따라서 계대에 따라 체세포의 염색체이상의 비율이 상대적으로 증가하고, 체세포 핵이식에 따른 염색체 이상이 생길 수 있음을 알 수 있었다. (이 논문은 농림부 연구비에 의해 수행되었음)

• Korean Journal of Plant Tissue Culture
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• v.24 no.3
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• pp.175-181
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• 1997

Plants were regenerated from hypocotyl protoplast culture of cabbage (F$_1$hybrid Green Challenger) and were transplanted on fields. The ploidy level of regenerated and seed-grown plants was measured by flow cytometry. In total 125 regenerated plant, diploid (2n), tetraploid(4n), and mixoploid (2n+4n) were 72.8%, 25.6%, and 1.6%, respectively. Most of the regenerated plants had normal phenotype, whereas several plane showed abnormal phenotype such as heavy leaf incision, savoy, and wave. The regenerated plants with abnormal phenotype and different ploidy level were analysed by isozyme and RAPD, but no significant difference was found.