• 한국발생생물학회지:발생과생식
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• 제17권4호
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• pp.345-351
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• 2013

This study examines the effects on fertilization rate (FR), hatching rate (HR), and normal individual rate after artificial fertilization using frozen thawed sperm according to the cryoprotectant (DMSO) concentration and the period of cryopreserved sperm of longtooth grouper, Epinephelus bruneus. Performing artificial fertilization using frozen-thawed sperm, after freezing the sperm at different DMSO concentration of 5.0%, 7.5%, 10.0% respectively, FR were (DMSO 5.0%: $99.5{\pm}0.8%$, DMSO 7.5%: $99.5{\pm}0.7%$, and DMSO 10.0%: $99.6{\pm}0.6%$). The results are not significantly different from the control fresh sperm (100%). HR also (DMSO 5.0%: $96.2{\pm}2.3%$, DMSO 7.5%: $95.3{\pm}3.6%$, 10.0%: $96.6{\pm}1.8%$) were not significantly different in each group. The normal individual rate after hatching using with control fresh sperm ($98.4%{\pm}0.5$) and DMSO concentration level of 5.0% ($97.8{\pm}0.1%$) were not significantly different. However, with 7.5% ($97.2{\pm}0.6%$) and 10.0% DMSO concentrations ($95.9{\pm}0.2%$) are lower than the normal individual rate after hatching observed in the control and 5.0% DMSO. Performing artificial fertilization using frozen-thawed sperm at different frozen period (2 days, 2 years, and 3 years), 10% DMSO FR and HR of 3 years (FR; $66.8{\pm}1.8%$, HR: $82.0{\pm}12.9%$) and 2 years (FR; $78.5{\pm}14.8%$, HR: $79.3{\pm}0.6%$) cryopreserved sperm were lower than control (FR; 100%, HR: $91.1{\pm}3.6%$) and 2 days cryopreserved sperm (FR; $99.6{\pm}0.6%$, HR: $96.6{\pm}1.8%$). These results suggest suitable DMSO concentration ranges of cryopreservation sperm for E. bruneus is 5 to 10% and with 2 to 3 years cryopreservation period, cryopreservation sperm can be useful for seed production.

• 한국양식학회지
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• 제17권3호
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• pp.180-186
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• 2004

본 연구는 동갈돗돔의 대량 인공종묘생산 기술개발을 위한 기초자료를 얻고자 자연산란을 유도하여 산란된 알의 특성을 조사하였다. 실험에 사용된 어미는 3년동안 실내사육한 자연산 7마리로서, 크기는 전장 34.0∼44.0 cm (38.6$\pm$4.0 cm), 체중 1.00∼2.23 kg (1.62$\pm$0.50 kg)이었다. 산란은 2000년도에는 9월 22일부터 10월 1일까지 9일간, 2001년도의 경우, 8월 22일부터 10월 3일까지 37일 동안 이루어졌다. 산란기간 중 수온은 19.8∼28.5$^{\circ}C$범위였다. 알의 ml당 수는 1,700개였고, 2년 간 총 산란량은 22,883,000개로, 이 중 부상률은 41.7% (9,539,000개)였다. 부상란에 대한 수정률은 85.0∼99.9%였으며, 부화율은 2.9∼93.0%였다. 수정란은 분리부성란으로 난경은 0.85∼0.98 mm였으며, 유구는 1∼5개가 존재하였다. 수온 26 $^{\circ}C$ 에서 수정 10분 후에 배반이 형성되었으며, 수정 후 50분에는 16세포기, 3시간 후에는 포배기, 7시간 30달 후에는 배체가 형성되었다. 부화는 수정 20시간 40달 후에 이루어졌으며,부화자어의 크기는 전장 1.81$\pm$0.18 mm이었다. 수온별 부화소요시간은 23$^{\circ}C$에서는 31∼34시간, 29$^{\circ}C$의 경우, 17∼20시간이 소요되었다.

• 대한수의학회지
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• 제38권2호
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• pp.394-400
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• 1998

Cryopreservation of embryos by vitrification is a simple method to preserve bovine embryos for subsequent embryo transfer, but embryonic viability after vitrification has been inconsistent and low compared with conventional slow freezing. The aim of the present study is to examine the effect of serum or serum albumin in a vitrification solution and epidermal growth factor(EGF) or fibroblast growth factor(FGF) on in vitro viability of bovine blastocysts frozen by vitrification. Bovine blastocysts were produced by in vitro maturation, fertilization of follicular oocytes and culture of embryos in a synthetic oviduct fluid medium(SOFM) containing BSA and 19 essential and nonessential amino acids. Blastocysts with excellent or good morphology were selected at 7 or 8 days after culture and utilized for vitrification. In experiment 1, blastocysts were vitrified in a solution containing semi-fetal calf serum(SFCS) or BSA(5 or 10mg/ml) and then their subsequent viabilities were examined by culturing thawed embryos in a SOFM containing BSA and 19 amino acids. Effect of EGF or FGF added to a SOFM containing polyvinyl alcohol(PVA) on the viability of vitrified-thawed blastocysts was investigated in experiment 2. BSA added at 5 or 10mg/ml to a vitrification solution showed significantly higher(p < 0.05) developmental rate to expanded and hatching blastocysts than SFCS, but there was no significant difference in the developmental rate to hatched blastocysts after thawing. Supplementation of a culture medium with EGF and/or FGF significantly increased(p < 0.05) embryo development to expanded blastocysts compared with control but showed no beneficial effect on the development to hatching or hatched blastocysts. Coculture of thawed embryos with granulosa cells in a TCM 199 containing 10% fetal calf serum(FCS) showed the highest developmental rate to expanded, hatching and hatched blastocysts among the groups tested. In conclusion, supplementation of a vitrification solution with BSA at 5mg/ml and culture of thawed blastocysts in a medium containing EGF and/or FGF can improve in vitro viability of bovine blastocysts frozen by vitrification.

• 한국수산과학회지
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• 제37권2호
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• pp.98-104
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• 2004

Early development and metamorphosis of the ascidian (Halocynthia hilgendorfi ritteri) were investigated from fertilized egg. The samples were collected in the coastal waters of Yongdam, northwest of Jeju Island in November 2002. H. hilgendorfi ritteri was solitary ascidian and produced spheral eggs with egg size ranging from $0.33\pm0.01\;mm.$ On the outer surface of the vitelline coat are attached many follicle cells. At $21.0\pm0.5^{\circ}C$ of water temperature, first cleavage took place in about 1.5 hrs after fertilization, and gastrulation followed in about 12.5 hrs. The formation of tailbud embryos and free swimming larvae were observed 13.3 hrs and 20.5 hrs after fertilization, respectively. The size of newly hatched tadpole larva was 1.30-1.45 mm, the larva swam for 2 hrs to 14 hrs. At 4 hrs after hatching, the palpi were lost and tail absorption began with an abrupt rupture of the anterior end of the notochord. At 17-18 hrs after hatching, tail completely absorption and remained trunk. The coniform adhesive papilla began protrusion at 30 hrs after hatching. The oral and atrial siphon formed at 6-7 days after settlement. At 17-18 days after settlement, metamorphosed the larvae developed into protoascidian of which the external morphology was similar to their adult.

• Clinical and Experimental Reproductive Medicine
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• 제20권1호
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• pp.9-17
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• 1993

This study was carried out investigate the effect of water quality and the kind of media on the in vitro development of 1-cell and stage mouse embryos. $F_1$ hybrid mice were superovulated and timely mated. 1-cell stage and 2-cell stage mouse embryos were recruited and taken into Ham's F-10 or m-KRB media which was made of two of two kinds of water having different quality, highly purified water and tap water. 2-cell stage embryos grew up well in vitro to blastocyst or hatching blastocyst regardless of the composition of culture media, but 1-cell stage mouse embryo didn't develop well to blastocyst or hatching blastocyst in simple media like m-KRB. These results meant in vitro devleopment of 1-cell stage mouse embryo neded complex media like Ham's F-10 which contained abundant protein components. In case of quality control for water, in vitro fertilization program. observation of in vitro development of 2-cell mouse embryos up to blastocyst or hatching blastocyst media such as m-KRB would be efficatious in detecting the difference of water quality.

• 한국수산과학회지
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• 제51권4호
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• pp.420-425
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• 2018

We characterized egg development and investigated the effects of temperature on embryonic development time, in the convict cichlid Amatitlania nigrofasciata. Fertilized eggs of this species have an ovoid shape (horizontal axis, $1.56{\pm}0.02mm$; vertical axis, $1.26{\pm}0.04mm$) and smooth translucent chorion surrounded by a layer of mucous secretion. At $27{\pm}0.5^{\circ}C$, the first cleavage, blastula, gastrula and 16-somite stages of A. nigrofasciata eggs began at 1.5, 4.83, 14 and 40 hours after fertilization, respectively. We measured the development rate of embryos at $12-33^{\circ}C$ and found that the time period from fertilization to hatching was 94 hours at $24^{\circ}C$, 64 hours at $27^{\circ}C$ and 50 hours at $30^{\circ}C$. At temperatures of $12-21^{\circ}C$, all fertilized eggs died before hatching and those incubated at $33^{\circ}C$ died immediately after hatching.

• Environmental Analysis Health and Toxicology
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• 제25권3호
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• pp.241-251
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• 2010

Crucian carp (Carassius auratus) has been used as the sentinel species for POPs (Persistent Organic Pollutants) monitoring in aquatic environment. However, there is little information for dioxin toxicity and especially, early life stage toxicity in crucian carp have been never carried out. In this study, we investigated several toxic effects for 2,3,7,8-tetrachlorodibenzo-p-dioxin (2,3,7,8-TCDD) in fertilized egg obtained by natural fertilization from crucian carp. The embryos at 3 h post-fertilization (hpf) were treated with 0.039, 0.156, 0.625, and 2.5 (${\mu}g/L$) TCDD by waterborne exposure for 60 minutes and changed with fresh water 2 times per day. Fertilized eggs started hatching at 51 hpf and TCDD exposed embryo showed decrease of hatching rate in a dose-dependent manner at 75 hpf. Pericardial edema was continuously observed in larvae exposed to TCDD from hatching start time (51 hpf), followed by the onset of mortality. In addition, AhR-related gene, CYP1A was clearly increased by TCDD in a dose dependent manner. These results indicated that fertilized eggs obtained from crucian carp have the TCDD related gene regulation and a distinct TCDD developmental toxicity syndrome by TCDD exposure. Therefore, we suggested that early life stage test in crucian carp could be used as test methods on dioxins toxicity.

• Ocean and Polar Research
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• 제42권4호
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• pp.303-311
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• 2020

In this study, the embryonic development and hatchability of eggs fertilized by the reciprocal hybrids of kelp grouper (Epinephelus bruneus) and red-spotted grouper (E. akaara) were evaluated with the goal of establishing a novel hybrid with enhanced growth and viability during the farming period in the temperate waters of Korea. The fertilization rates were lower for hybrids than for maternal purebreds and were significantly higher in the red-spotted grouper ♀ × kelp grouper ♂ hybrid (RGKG, 89.61 ± 1.58%) than in the kelp grouper ♀ × red-spotted grouper ♂ hybrid (KGRG, 74.82 ± 4.23%, p < 0.05). Unlike the fertilization rates, the hatching rates of fertilized eggs were similar between hybrids and maternal purebreds and did not differ significantly between KGRG and RGKG (72.74 ± 3.60% vs. 75.23 ± 2.20%, respectively, p > 0.05). The embryonic development of the hybrids was similar to that of maternal purebreds; however, irregular cleavage and asymmetric blastoderm were noticeable in the developing eggs of KGRG hybrids. The deformity rates of newly hatched larvae were higher in hybrids than in maternal purebreds and were significantly higher in KGRG than in RGKG (17.47 ± 1.28% vs. 7.11 ± 0.54%, respectively, p < 0.05). These results demonstrate the potential to produce viable larvae from these two hybrids. Although the production efficiency of KGRG was lower than that of RGKG, the fertilization, hatching, and deformity rates make both hybrids useful for further comparative studies regarding economic aspects.

• 한국패류학회지
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• 제31권3호
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• pp.179-186
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• 2015

굴 미수정란의 동결보존에 적합한 동해방지제의 종류, 농도 및 침투 시간을 구명하기 위하여 4가지 동해방지제 (DMSO, EG, methanol, PEG) 를 대상으로 수정률, 부화율 및 형태 이상률을 조사하였다. 동해방지제의 농도 및 침지시간은 각각 5, 10, 15, 20%와 10, 20, 30, 40분이었다. 침지 후 미수정란은 정상 정자와 수정되었고, 수정률, 부화율 및 형태 이상률은 동해방지제의 종류와 농도 및 침지시간에 영향을 받았다. 동해방지제의 농도와 침지시간의 증가에 따라 수정률과 부화율은 낮아졌고, 대부분의 동해방지제에서 5-10% 농도와 10-20분의 짧은 침지시간에서 좋은 결과를 보였다. 수정률과 부화율을 근거로 굴 미수정란에 가장 적합한 동해방지제는 DMSO였으며, 처리 농도와 침지 시간은 각각 5%와 10-20분이었다.

• 한국양식학회지
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• 제12권1호
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• pp.39-45
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• 1999

해삼의 인공종묘생산 기술개발을 위한 생물학적 기초자료로 이용하고자 산란유발, 유생발달 과정, 유생 사육시 먹이와수용밀도에 관한 시험을 실시한 결과는 다음과 같다. 산란유발은 수온자극에서 6.0~17.5%의 반응이 있었으며, 간출자극에서는 1.4~4.0%로 낮은 반응률을 보였다. 산란량은 50~500만개였고, 수정률은 84.0~96.0%였으며, 부화율은 71.4~84.6%로 나타났다. 해삼의 수정난은 $154{\mu}m$이며, 난발생은 수온 $23^{circ)C$에서 수정후 2시간 10분만에 4세포기, 3시간 40분이 지나면 16세포기로 되며, 14시간 30분 후에는 포배기로 되면서 부화하였다. 그 이후 3일째에는 auricularia 유생으로 되었고, 11일째에는 doliolaria 유생, 15일째에 pentactula 유생으로 변태하였다. 유생의 먹이로는 Chaetoceros calcitrans와 Pav-lova lutheri가 적당한 것으로 나타났으나, 이 중 Chaetoceros calcitrans는 변태가 빠르고 생존율도 높아 해삼 유생의 먹이로 가장 좋았으며, 유생 사육을 위한 최적 밀도는 2개체/ml 이하를 유지하는 것이 좋은 것으로 나타났다.