• Journal of Aquaculture
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• v.17 no.2
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• pp.97-102
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• 2004

We examined the effects of stimulants including sunlight and UV-irradiation on the spawning induction and early development of the noble scallop, Chlamys nobilis. The sunlight stimulation resulted in nuch faster spawning induction (100% success within 40 minutes) compared to UV-irradiation (100% success within 70 minutes). Early development of the scallop larva took place between 15$^{\circ}C$ to 3$0^{\circ}C$. The time to reach the early D-shaped stage was 63.5, 31.5, 18.5 and 17.0 hours at 15, 20, 25 and 3$0^{\circ}C$, respectively. The correlations between the water temperature-(WT) regimes and the time (t) required for each developmental stage are as follows. 2 cell stage: 1/t=0.0606WT-0.6194 ($r^2$=0.9791) 8 cell stage: 1/t=0.0304WT-0.3453 ($r^2$=0.9941) Morula: 1/t=0.0100WT-0.1049 ($r^2$=0.9663) Trochophore: 1/t=0.0058WT-0.0618 ($r^2$=0.9848) D-shaped larva: 1/t=0.0030WT-0.0282 ($r^2$=0.9731) These correlations indicated that the biological minimum temperature of the species is around 10.44$^{\circ}C$. The highest survival rate up to D-shaped larva at different water temperature was observed at $25^{\circ}C$.

• Clinical and Experimental Reproductive Medicine
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• v.28 no.1
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• pp.41-46
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• 2001

Objective: To determine the effects of leukemia inhibitory factor (LIF) on embryonal development in in vitro culture. Methods: This is designed in vitro model using eggs from mouse. The eggs from mouse were assigned 29 for control group, 53 for 20 ng/ml of LIF, 88 for 40 ng/ml of LIF, 68 for 80 ng/ml of LIF respectively for in vitro fertilization. And 26 fertilized eggs at 2 cell stage from mouse also were assigned. The mouse embryos of all groups were cultured in medium supplemented with LIF in different concentrations, whereas the eggs in control group was cultured in medium without supplement of LIF. Results: At 72 hours culture of eggs from in vitro fertilization, there was a slight increas in rate of embryonal development to morula in both LIF-20 and LIF-40 as results of 64.15% and 75% respectively, while 42.65% in inferior rate of LIF-80, compare with 51.72% in control group. But the difference between these each groups were not significant in statistically ($p{\le}0.05$). And after 96 hours culture of eggs, the rates blastocyst formation was significantly higher in both LIF-20 and LIF-40 as 56.6% and 63.63% than those in control and LIF-80 as 44.83% and 35.29% respectively. On culturing eggs from in vivo fertilization, the rates of blastocyst formation was significantly not only higher as 85% and 81.81% respectively in medium supplemented with LIF-40 and LIF-80 than 42.3% in LIF-20 but also embryonal cell viability were remakedly improved at 96 hours after culture. Conclusion: The LIF in low dose is embryotrophic, but LIF in high dose is embryotoxic on eggs from in vitro fertilization. Whereas on culturing eggs from in vivo fertilization, LIF is more beneficial with dose dependent in high concentration.

• Journal of Embryo Transfer
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• v.10 no.2
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• pp.105-113
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• 1995

This study was conducted to examine the efficiency of enucleation and blastomere isolation from recipient oocytes and donor embryos, respectively and to determine the effect of oocyte age and electric voltage on the fusion rate and in vitro development of the fused oocytes in rabbit nuclear transplantation. Immature oocytes collected from ovarian follicles were matured in vivo for 12 h in TCM-199 containing FCS and hormones and in vivo matured oocytes were collected 17 to 18 h post-HCG. The fresh and frozen donor embryos of 8- to 16-cell stage were collected from the oviduct of superovulated does. The proportion of successfully enucleated oocytes was greatly lower in in vitro matured oocytes (42.3%) than that (62.7%) in in vivo matured oocytes The level of cytochalasin B for in vivo matured oocytes did not affect the efficiency of enuleation, but 7.5 $\mu$g /mL cytochalasin B for in vitro matured oocytes showed a high enucleation rate significantly. The isolation efficiency of a single blastomere nucleus did not differ between 8- and 16-cell stage embryos. The percentage of single blastomeres isolated from 16-cell stage fresh embryos after 0.5% pronase treatment was greatly higher at 16-min treatment (94.4%) than at 8-min(78. 1%) and the blastomeres(61.5%) isolated from frozen-thawed embryos after 16-min pronase were significantly fewer than those of fresh embryos. The age of recipient oocytes affected nuclear fusion rate. The reconstituted oocytes fused at 24-h age showed slightly higher fusion rate (77.8%) than those (65.0%)fused at 18-h age. The fusion rate of in vitro and in vivo matured oocytes inserted with fresh blastomere did not differ among electric voltages, but the cleavage rate and development to morula-blastocysts of in vitro matured oocytes was more higher under 0.6 kV/cm than under 0.8 to 1.2 kV/cm, while the cleavage rate and development of in vivo matured oocytes was higher under 0.8 to 1.0 kV/cm than under 1.2 kV/cm. The fusion and cleavage rate fol1owing insertion with frozen-thawed blastomere was not different between the in vitro and in vivo matured oocytes and was similar to those from fresh blastomere insertion.

• Journal of Embryo Transfer
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• v.19 no.3
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• pp.257-264
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• 2004

For safe preservation of Korean Native Pigs (KNP) as an animal genetic resources and a means to maintain the genetic diversity, we performed to investigate the optimal hormone levels for superovulated gilts and establish the cryopresevation methods of embryos. The reults were as follows; 1. The number of ovulated corpus luteums (CL) and follicles were 12.4, 13.6, 30.0 or 23.3 in hCG 500IU and PMSG 500, 750, 1,000IU or hCG 750IU and PMSG 1,000IU respectively. In the case of PMSG 1,000IU와 hCG 500IU, there showed highest number but were no significance among others. The recovery rate of embryos by the ovulated CL were 59.4-79.2％. 2. The morula stage embryos recovered on Day 4 after insemination were significantly higher than Day 5 (P<0.01), but blastocyst stage embryos recovered on Day 5 were sinificantly higher than Day 4 (P<0.05). 3. The survival rate of expanded blastocyst were 23.5％ in conventional freezing with 1.4 M glycerol.

• Korean Journal of Ichthyology
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• v.10 no.2
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• pp.184-190
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• 1998

The early life history of black bullhead, Pseudobagrus koreanus, endemic to Korea was investigated to get biological information needed in artificial production of seedlings and in recovering natural resources. The fertilized eggs showed some characteristics in having heavy sticky material and minute folds which is formed radical pattern on the egg membrane. The shape of egg was spherical and $2.59{\pm}0.08$(2.45~2.70, n=10)mm in diameter. The yolk had not oil globule. The first cleavage was observed 2 hrs after insemination at $21{\sim}23^{\circ}C$, and the progressive cleavage were done about 30 min. interval. The characteristic changing of the yolk surface started at morula stage and continued to the end of gastrula. Hatching was started 72 hrs and completed 90 hrs after fertilization. The size of the larvae were 5.41~5.81mm in total length and 2.76~2.94mm in preanal length, and the number of so mites was 15-16+33~34(48~50). The barbels and swimbladder were completed and all the fins except second dorsal were appeared 1 week after hatching. The larvae attained 9.67~10.52mm in total length and 5.20~5.65mm in preanal length. All the fin sets and color pattern were completed 2 weeks after hatching and body mucus was secreted at that stage. The juvenile attained 14.59~16.02mm in standard length, 3.31~4.16mm in head length and 8.07~9.31mm in prenal length.

• Asian-Australasian Journal of Animal Sciences
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• v.15 no.12
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• pp.1695-1701
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• 2002

The purposes of this study were to optimize the in vitro maturation (IVM) and culture (IVC) systems of rabbit oocytes. Cytoskeletal structures in the germinal vesicle stage (GV) and during IVM are also investigated. Ovaries were transported from local slaughterhouses and the cumulus-oocyte complexes (COCs) were collected from ovarian follicles (${\geq}1mm$). COCs were randomly allocated to TCM199-based medium ($T_1$, TCM-199) supplemented with $NaHCO_3$, glucose, sodium pyruvate and FSH ($T_2$), $T_2+E_2+LH$ ($T_3$), $T_3+FBS$ ($T_4$), or $T_1+E_2+LH+FSH+FBS$ ($T_5$), for IVM. In Experiment 1, COCs were retrieved from the follicles and 51 GV oocytes were fixed in the fixative (MTSB-XF) for nuclear and cytoplasmic examinations. In Experiment 2, progressive changes of both the nucleus and the cytoskeleton were examined at 0, 6, 16, and 20 h after IVM. Maturation (MR) and developmental rates were assessed in Experiment 3. Cytoplasmic microtubules (MT) were clearly observed in rabbit GV oocytes. To our knowledge, this is the first report that describes the appearance of MT structures in the GV stage ooplasm. Tremendous variations in cytoskeletal alterations were observed among treatments with the exception of the vitelline ring (VR), which is constantly visible and unchanged during maturation. Germinal vesicle breakdown (GVBD) does not occur at 6 h after onset of maturation culture. When the oocytes for IVM were collected within 2 h, results from Experiment 3 showed that rates of nuclear maturation were 42, 8, 42, 37 and 65% at 16 h of IVM for $T_1$ through $T_5$, respectively, in which $T_1$, $T_4$ and $T_5$ had significantly greater MR than those in other groups (p<0.05). Morula/blastocyst development after parthenogenetic activation ranged from 20 to 63% with significantly greater rates in $T_3$, $T_4$ and $T_5$ (p<0.05). These results suggested that oocytes recovered from slaughterhouse ovaries can be matured and parthenogenetically activated in vitro, but the MR remained low in this study. Addition of $E_2$ and LH in the medium may be beneficial for cytoplasmic maturation, but FBS exerts a nega- tive role in the subsequent development of parthenogenetic embryos when energy substrates are provided in the IVC media. More studies are required for improving the MR and further development of the GV stage rabbit oocytes.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.11 no.2
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• pp.115-121
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• 1978

Early developmental stages of Mactra chinensis were bioassayied to determin the water quality of the costal waters of Busan. The water samples were obtained at 12 stations from July 12 to July 17, 1977. Water quality evaluation was made in terms of the rates of normal development to abnormal development in three indicatory states, i. e. fertilization membrane formation, morula and trochophore larvae. The pollution degree of the waters brought from five swimming beaches was found to be highest at Haewundae followed by Songdo, Kwang-anri, Songjeong and Dadaepo with decreasing order. The highest value of the water at Haewundae was mainly due to the accidental oil spill from an oil tanker which happened on July 7, 1977. At Dongsamdong the rate of total abnormal development was $47.17\%$. This result is apparently attributable to fish byproducts discarded from a number of restaurants. At Chungmudong where a large fish-market is located, the rate of total abnormal development was $61.14\%$. At Suyeng Bay and Yongho Bay the rates of total abnormal development were $73.82\%$ and $72.90\%$ respectively. At these bays the drainage presumably contains a large amount of chemical pollutants from the industrial areas. This result shows that no organisms can normally breed in these regions.

• Ocean and Polar Research
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• v.40 no.3
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• pp.161-167
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• 2018

We examined the effect of salinity and water temperature on hatching and survival rates of fertilized eggs of hybrid grouper (Epinephelus fuscoguttatus ♀ ${\times}$ E. lanceolatus ♂) at different developmental stages, determining optimal conditions for their long-distance transportation. Deformities and hatching rates of fertilized grouper eggs were observed at salinities of 24, 27, 30, 33, 36, and 39 psu. The optimal salinity was determined to be 36 psu, with a survival rate of $70.0{\pm}2.0%$. Transportation experiments at 36 psu were conducted at water temperatures of 21, 24, 27, and $30^{\circ}C$, different developmental stages such as morula, 5-myomere, and tail beating for hatching and survival rates. The optimal water temperature and developmental stage for transporatation were $30^{\circ}C$ and tail beating stage and those hatching rates were $50.6{\pm}1.9%$ and $86.3{\pm}1.3%$, respectively. At $21^{\circ}C$, the survival rate by transportation water temperature was highest ($73.1{\pm}10.6%$), but the hatching rate ($17.1{\pm}3.1%$) was lowest. Therefore, the hybrid grouper fertilized eggs (E. fuscoguttatus ♀ ${\times}$ E. lanceolatus ♂) can be most efficiently produced under long-distance transportation conditions during the tail beating stage and at a water temperature of $30^{\circ}C$.

• Korean Journal of Animal Reproduction
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• v.14 no.2
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• pp.101-106
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• 1990

These mxperiments were carried out to investigate the effect of rabbit anti-bovine cumulus cell antibodies on in vitro fertilization and following development of bovine follicular oocytes matured in vitro. The bovine ovaries were obtained at a slaughter house and the follicular oocytes surrounded by cumulus cells were collected by puncturing follicles with 2~6mm of diameter. Bovine oocytes were matured in vitro for 24~26hrs in a CO2 incubator with 5% CO2 in air at 39$^{\circ}C$ and subsequently cultured in medium containing cumulus cell antibody for 1 hour. The medium used for maturation was TCM-199 supplemented with hormones, pyruvate, FCS and antibiotics. Epididymal spermatozoa were capacitated by in vitro culture for 2~3 hrs in BO solution 10~15 matured oocytes into the suspension of capacitated spermatozoa. Six hour after insemination the eggs were transferred to TCM-199 supplemented with FCS(10%) and then cultured for 7 days. The results obtained in these experiments were summarized as follows : 1. When the follicular oocytes matured in vitro were treated with antibody to intact cumulus cells, the fertilization rate of cumulus intact and removed oocytes was ranged to 45.0 to 53.7%. These value is slightly lower than that(64.3%) of follicular oocytes not treated with the antibody, and increased frequency of both male and female pronuclear formation was found in cumulus intact oocytes cultred in medium without the antibody(p<0.05). 2. The fertilization rate of cumulus intact and removed oocytes treated with antibody to solubilized cumulus cells was ranged 45.0 to 52.5%, significantly lowre than that(62.8%) of oocytes cultured in antibody free medium, and increased frequency of ova with male and female pronuclei was found when cumulus cells were present(p<0.05). 3. The rates of cumulus cell intact and removed oocytes developed to 8-, 16-cell and morula or blastocyst after treatment of intact and solubilized cumulus cell antibody were ranged 7.1 to 14.5, 2.9 to 5.9 and 1.5 to 2.9%, respectively, slightly lower than 18.6, 10.0 and 8.6% of cumulus intact oocytes cultured in medium without the antibody. The results of this stduy indicate that cumulus cells promote not only normal fertilization with proper pronuclear formation, but embryo development and that the beneficial effect of cumulus cell to the pronuclear formation and embryo development is blocked by the action of antibody to cumulus cell.

• Korean Journal of Animal Reproduction
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• v.18 no.1
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• pp.55-62
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• 1994

This study was carried out to test the effects of fluorescein diacetate(FDA 0${mu}ell$/ml, 0.5${mu}ell$/ml, 5${mu}ell$/ml, 10${mu}ell$/ml or 0${mu}ell$/ml, 5${mu}ell$/ml in PBS) treated before culture on the survival of vitrified mouse morulae in vitrification solution(20% glycerol＋glycerol＋10% ethylene glycol＋30% ficoll＋10% sucrose). The results were summarized as follows; 1. The survival rate of FDA-tested fresh mouse morulae after 24 hours culture was over 96%((P4.8) in the control or treatment groups with various levels of FDA. Because the rate of mouse morulae which developed to hatched blastocysts was higher with the various levels of FDA treatment(67%) than control(50%), it was considered that toxicity of FDA did not affect the survival of mouse morulae. 2. When mouse morulae were FDA-tested in FDA 0(control), 0.5, 5, and 10${mu}ell$/ml treatment after vitrification, the development rate to expanded blastocyst were 66, 82, 64 and 76%, and FDA scores were P4.2(84%), P4.7(94%), P4.2(84%) and P3.9(78%), respectively. There were no significant differences between control and FDA treatments, but there were significant difference between 0.5${mu}ell$/ml)94%) and 10${mu}ell$/ml(78%) treatment(P<0.01). 3. The survival rates of cultured mouse morulae according to FDA-scores(P0=non-fluorescence; P1~P5=according to their fluorescence) after vitrification were P5;92%, P4;67%, P3;42% and P2.P0;0%, respectively. 4. Implantation rates of morulae stage embryos cultured into early blastocysts and implanted into uterine hornes vitrification were 14 and 11% embryos treated control(0${mu}ell$/ml) and FDA 5${mu}ell$/ml and the normal fetus development was 2% embryos for both treatments. Results of this percent study indicated that toxicity of FDA does not affect not only the survival of fresh and vitrified mouse morulae but also the development rate and implantation of fetus after transfer as well. The development rates of mouse morulae with the FDA score of P5, P4 and P3 were 92, 67 and 42%, respectively, it was considered that FDA-test was fit for the judge of survival.