• Korean Journal of Animal Reproduction
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• v.20 no.1
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• pp.77-84
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• 1996

The purpose of this study was to evaluate effects of osmolarity and vitamins on the in vitro development of embryos. Bovine embryo that had been matured and fertilized in vitro were cultured in simple, chemically defined, protein free medium (mTLP-PVA). When the osmolarity of the medium supplemented 0.35 mM phos-phate and 19 amino acid was changed by NaCI concentration, significantly(p<0.05) higher pro-portion of 1-cell embryos in the medium of 265 or 290 mOsm developed to the morula (32 ~ 35%) and blastocyst(24 ~ 28%) stage. When embryos were transferred to fresh medium containing 5.56mM glucose at 120hrs post-insemination, the highest proportion of embryos developed to mor-ula(40%) and blastocyst(32%) stages at 290 mOsm(p<0.05), although the value in morulae was not significantly different with that(35%) at 315 mOsm. Vitamins in presence of glutamine and amino acids had no beneficial effects on the development of 1-cell embryos to the blastocyst.

• Asian-Australasian Journal of Animal Sciences
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• v.20 no.6
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• pp.842-849
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• 2007

The purpose of the present study was to produce live offspring from in vitro fertilized goat embryos. Oocytes were collected from abattoir ovaries and kept in oocyte collection medium. Oocytes were washed 4-5 times with maturation medium containing medium-199 with 5 ${\mu}g/ml$ FSH, 100 ${\mu}g/ml$ LH, 1 ${\mu}g/ml$ estradiol-$17{\beta}$ 50 ${\mu}g/ml$ gentamycin, 10% inactivated estrus goat serum, and 3% BSA (fatty acid free). Oocytes were placed in 100 ${\mu}l$ drops of maturation medium containing granulosa cell monolayer and incubated in a 5% $CO_2$ incubator at $38.5^{\circ}C$ for 27 h. For capacitation of spermatozoa fresh semen was processed and mixed in 3 ml fertilization TALP medium containing 50 ${\mu}g/ml$ heparin and kept in the above incubator for 2 h. The capacitated spermatozoa were coincubated with matured oocytes for fertilization. Cleaved embryos were separated and cultured in embryo development medium with oviductal cells and 494 embryos were produced. Recipient goats were synchronized with two injections of 15 mg $PGF_{{2}{\alpha}}$/goat 10 days apart. Eighty early stage embryos were transferred into the uterotubal junction of 14 surrogate mothers using laparoscopy techniques. One recipient delivered twin kids, whereas another two recipients each.delivered a single kid The parentage of these kids was evaluated using highly polymorphic co-dominant microsatellites markers. From the present study, it was concluded that live goat kids can be produced from in vitro matured and fertilized goat embryos, to the best of our knowledge for the first time in India.

• Clinical and Experimental Reproductive Medicine
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• v.47 no.3
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• pp.221-226
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• 2020

Objective: We attempted to identify the optimal cutoff numbers of mature oocytes that would produce at least one or multiple top-quality (grade A) day-3 embryos in normal responders undergoing stimulated in vitro fertilization (IVF) cycles. Methods: We selected 210 fresh IVF cycles performed in 170 infertile women at a single center from January 2014 to November 2019. Four to 14 (total) oocytes were obtained in all cycles after conventional ovarian stimulation. A receiver operating characteristic curve analysis was performed to find the moderate and extreme cutoff numbers of mature oocytes that would produce ≥ 1, ≥ 2, ≥ 3, ≥ 4, and ≥ 5 top-quality embryos. Results: The cutoff number of mature oocytes was significantly correlated with the number of top-quality embryos (r = 0.467, p= 0.000). The moderate cutoff number of mature oocytes was ≥ 3, ≥ 5, ≥ 5, ≥ 6, and ≥ 6 for obtaining ≥ 1, ≥ 2, ≥ 3, ≥ 4, and ≥ 5 top-quality embryos, respectively. The extreme cutoff number of mature oocytes was ≥ 9, ≥ 9, ≥ 10, ≥ 10, and ≥ 11 for obtaining ≥ 1, ≥ 2, ≥ 3, ≥ 4, and ≥ 5 top-quality embryos, respectively. Conclusion: We present the optimal cutoff numbers of mature oocytes that would yield ≥ 1, ≥ 2, ≥ 3, ≥ 4, and ≥ 5 top-quality embryos with 95% specificity. Our findings could help infertility clinicians to set target mature oocyte numbers in women undergoing stimulated IVF cycles.

• Proceedings of the Plant Resources Society of Korea Conference
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• 2000.10a
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• pp.114-115
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• 2000

Aralia elata is found in mountain areas all over Korean peninsula. Aralia elata is the scientific name for Japanese angelica tree. The tree belongs to the family Araliaceae, commonly known as ginseng family. Bud sprouts from apical shoot tip of the plants are rich in flavor and thus mainly used for both folk medicine and vegetable. The stalks with apical buds are gathered in the early spring and planted in sandy soil or water in the greenhouse. The sprouting buds are then collected and sold as fresh vegetable. Although the plants have been used for food, they have been cultivated in a very small scale. In spring, local farmers just go around mountain areas to search the trees and gather the stalks as much as they get and sell them to the market. No conservation efforts have been made to stop the exploitation or to save the dwindling population. We tried to provide local farmers with the plants that may be used as an alternative to stalks from wild populations. This will bel! p conserve the wild populations. However, it is hard to propagate them either by conventional cuttings or by seed germination in a short period of time. Mass propagation using tissue culture systems have shown a great promise with several woody plants. Recently we developed a mass propagation technique via somatic embryogenesis system using mature and/or juvenile explants for Aralia elata. Several factors affecting somatic embryogenesis system including SE(somatic embryo) induction, embryogenic callus proliferation, SE germination, plant regeneration and transplanting to field frill be presented. And some problems arising for the somatic embryogenesis system will be also discussed.

• Clinical and Experimental Reproductive Medicine
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• v.38 no.3
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• pp.126-134
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• 2011

Preimplantation genetic diagnosis (PGD) is gradually widely used in prevention of gene diseases and chromosomal abnormalities. Much improvement has been achieved in biopsy technique and molecular diagnosis. Blastocyst biopsy can increase diagnostic accuracy and reduce allele dropout. It is cost-effective and currently plays an important role. Whole genome amplification permits subsequent individual detection of multiple gene loci and screening all 23 pairs of chromosomes. For PGD of chromosomal translocation, fluorescence $in-situ$ hybridization (FISH) is traditionally used, but with technical difficulty. Array comparative genomic hybridization (CGH) can detect translocation and 23 pairs of chromosomes that may replace FISH. Single nucleotide polymorphisms array with haplotyping can further distinguish between normal chromosomes and balanced translocation. PGD may shorten time to conceive and reduce miscarriage for patients with chromosomal translocation. PGD has a potential value for mitochondrial diseases. Preimplantation genetic haplotyping has been applied for unknown mutation sites of single gene disease. Preimplantation genetic screening (PGS) using limited FISH probes in the cleavage-stage embryo did not increase live birth rates for patients with advanced maternal age, unexplained recurrent abortions, and repeated implantation failure. Polar body and blastocyst biopsy may circumvent the problem of mosaicism. PGS using blastocyst biopsy and array CGH is encouraging and merit further studies. Cryopreservation of biopsied blastocysts instead of fresh transfer permits sufficient time for transportation and genetic analysis. Cryopreservation of embryos may avoid ovarian hyperstimulation syndrome and possible suboptimal endometrium.

• Proceedings of the Plant Resources Society of Korea Conference
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• 2000.10b
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• pp.16-17
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• 2000

Aralia elata is found in mountain areas all over Korean peninsula. Aralia elata is the scientific name for Japanese angelica tree. The tree belongs to the family Araliaceae, commonly known as ginseng family. Bud sprouts from apical shoot tip of the plants are rich in flavor and thus mainly used for both folk medicine and vegetable. The stalks with apical buds are gathered in the early spring and planted in sandy soil or water in the greenhouse. The sprouting buds are then collected and sold as fresh vegetable. Although the plants have been used for food, they have been cultivated in a very small scale. In spring, local farmers just go around mountain areas to search the trees and gather the stalks as much as they get and sell them to the market. No conservation efforts have been made to stop the exploitation or to save the dwindling population. We tried to provide local farmers with the plants that may be used as an alternative to stalks from wild populations. This will hel! p conserve the wild populations. However, it is hard to propagate them either by conventional cuttings or by seed germination in a short period of time. Mass propagation using tissue culture systems have shown a great promise with several woody plants. Recently we developed a mass propagation technique via somatic embryogenesis system using mature and/ or juvenile explants for Aralia elata. Several factors affecting somatic embryogenesis system including SE(somatic embryo) induction, embryogenic callus proliferation, SE germination, plant regeneration and transplanting to field will be presented. And some problems arising for the somatic embryogenesis system will be also discussed.lso discussed.

• Environmental Analysis Health and Toxicology
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• v.25 no.3
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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.

• Korean Journal of Veterinary Research
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• v.35 no.3
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• pp.635-641
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• 1995

The purpose of this study was to investigate the effects of developmental stage and equilibration time on survival of rabbit embryos following freezing by vitrification. Adult New Zealand White female rabbits were superovulated with PMSG and hCG. The 8-cell stage embryos were collected from 40 to 45 hours after hCG injection by flushing oviducts with Dulbecco's phosphated buffered saline and in vitro cultured in TCM-199 containing 10% fetal calf serum(FCS). Each embryos developed in vitro to 16-cell, compact morula and blastocyst was cryopreserved and cultured following thawing to examine their developmental potential to expanded blastocyst stage in vitro. The frozen-thawed-cultured embryos were stained with Hoechst 33342, and their nuclei were counted using a fluorescence microscope. On the toxicity test of EFS solution as cryopreservation, the survival rates of 8-cell stage embryos was decreased in reverse to increasing of exposure time over 5 minutes. The post-thaw survival rates of embryos on equilibration times was significantly(P<0.05) higher for 2 min. than for 5 or 10 minutes. From morula to blastocyst of rabbit embryos was more suitable than 8-cell stage for cryopreservation by vitrification. The higher post-thaw survival rate of embryos can be achieved by keeping the cryoprotectant at $4^{\circ}C$ than at $20^{\circ}C$. The mean number of nuclei per embryo following freezing by vitrification and in vitro culture to expanded blastocyst at compacted morula and blastcyst was not significantly differ from fresh blastocyst.

• Forest Science and Technology
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• v.14 no.4
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• pp.160-168
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• 2018

The seeds of Elaeocarpus serratus, a tropical underutilized fruit tree are characterized by hard seed coat and consequent poor water uptake and low germination. To improve the regeneration through seeds, various parameters such as viability of seeds, water uptake, and effect of seed mass on germination and pretreatments were performed using a completely randomized design (CRD). Tetrazolium (TZ) test was conducted using fresh, mature seeds revealed $50{\pm}2.56%$ mean viability. Seeds of different weight classes showed similar pattern of water uptake and the saturation level was achieved at 60 hrs of soaking. Seeds belong to weight class 2.6-3.5g were germinated ($12.5{\pm}1.26%$) with $175{\pm}1.75days$ (d) of mean time taken for germination (MTG). Germination capacity of seeds varied significantly among different populations and Varkala population gave $12.5{\pm}1.1%$ germination with $174.6{\pm}2.5d$ MTG. Among various seed treatments, mechanical scarification was superior in germination and significant reduction in MTG ($p{\leq}0.05$). The mechanical scarification by complete removal of seed coat resulted in $49.2{\pm}1.52%$ germination within a short period of time ($9.52{\pm}0.89d$ MTG). However, the complete removal of seed coat without damaging to embryo is a difficult task. An alternate treatment (Mechanical scarification II) by making cracks on nut faces vertically followed by soaking in distilled water for 24 hrs gave $48.4{\pm}1.73%$ germination with significantly reduced MTG ($12.14{\pm}0.56d$) over unsoaked, untreated control ($6.5{\pm}1.84%$ germination and $197.18{\pm}1.79d$ MTG; $p{\leq}0.05$). This treatment (Mechanical scarification II) is therefore recommended for E. serratus seeds as it can adopt easily and can achieve 7 fold increases in germination over control. The recorded germination through mechanical scarification is in tune with realized viability percentage of the seeds.

• Journal of Embryo Transfer
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• v.19 no.2
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• pp.155-163
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• 2004

The aims of this study are 1) to test oocytes and embryos collected from in-vitro to achieving the valuable protocol by culturing, vitrifying and thawing of oocytes/embryos, and 2) to transfer them to recipient, and finally have resulted in pregnancies from recipient females after surgical or nonsurgical transfer. In vitro maturation and fertilization were performed according to Funahashi et al (1994). Glucose-free NCSU 23 supplemented with 5 mM sodium pyruvate, 0.5 mM sodium lactate and 4 mg/ml bovine serum albumin for 2 days at $39^{\circ}C$, and 10% fetal bovine serum albumin was added to the culture medium thereafter. Embryos were treated with 7.5 ${\mu}g/ml$ cytochalasin-B for 30 min, centrifuged at 13,000 rpm for 13 min and then exposed sequentially to an ethylene glycol(EG) vitrification solution, aspirated into OPS, and plunged/thawed into/from liquid nitrogen. In vivo embryos were surgically collected from three dornors after AI for control group. Forty-nine embryos were washed 3 times in mPBS + 10% FBS, followed treatments : cultured, centrifuged, vitrified, recovered and transferred to recipients as in vitro prepared embryos. Three recipients were transferred individually with 100, 100 frozen embryos derived from abattoir and 34 fresh embryos by surgically, and another three recipients were transferred individually with 150, 150 frozen embryos and 100 fresh embryos by nonsurgically, respectively. all recipient sows exhibited delayed returns to estrus. To our knowledge, theses results suggest that required an improved techniques, more vigorous embryos preparation and substitute to gilt with cleaner uterous condition.