• Fisheries and Aquatic Sciences
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• v.18 no.4
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• pp.417-420
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• 2015

We investigated the relationship between viability and IGF-1 receptor (IGF-1R) activity in D-shaped and umbo larvae of the surf clam Spisula sachalinensis after treatment with vitrification solution (VS) or freezing. In a toxicity assay, VS1, containing 5 M dimethyl sulfoxide (DMSO), was very harmful to D-shaped and umbo larvae. However, VS2, containing 5 M ethylene glycol (EG), was not harmful to either larval stage. Although VS2 had a promising toxicity test outcome, none of the larvae survived vitrification. After immersion into VSs and freezing, IGF-1R ${\beta}$-subunits were detected in all larvae; however, tyrosine phosphorylation of intracellular ${\beta}$-subunits was detected only in the control and live groups. These results suggest that activation of IGF-1R may influence surf clam larvae viability.

• Proceedings of the Korean Society of Developmental Biology Conference
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• 2003.10a
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• pp.72-72
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• 2003

The present study examined the possibility of cryopreservation of the D-shaped and umbo larvae of arkshell (Scapharca broughtonii), in terms of the survival rates after freezing and thawing. D-shaped and umbo larvae of arkshells were obtained from a shellfish farming on Yosu city. The average shell lengths were $93.3 \pm 10.1 \mu$m and $201.7 \pm 13.5 \mu$, respectively. Five cryoprotectants (CPAs), dimethyl sulfoxide (DMSO), glycerol, ethylene glycol (EG), propylene glycol (PG), and methanol, were tested at the concentrations of 1.5, 2.0 and 2.5 M. After larvae suspended in CPAs, cryoprotectants were loaded in 0.5 ml straws at a larval density of 50-100 larvae per straw, and epuilibrated for 10 and 20 minute at room temperature ($23^{\circ}C$), repectively. Straws were cooled at a rate of $1^{\circ}C$/min from $0^{\circ}C$ to $-12^{\circ}C$, held for 5 min at $-12^{\circ}C$, and then cooled at $2^{\circ}C$/min to $-35^{\circ}C$ and equilibrated for 5 min followed by plunging in liquid nitrogen. After storage in liquid nitrogen for 1 day, straws were thawed in a $30^{\circ}C$ water. As soon as straws were observed to melt, larvae were diluted with an equal volume of ASW and then washed twice with a large volume of ASW at an interval of 2 min to unload the CPAs. The results showed that after equilibration for 10 and 20 minute at room temperature, no larvae survived using methanol as CPAs, and it was observed that larval shells all open slightly, and larval flesh broke down and slopped over the shells. The highest survival rates (D-shaped larvae: 77.6%, umbo larvae: 59.3%) were obtained with 2M DMSO, and 1.5M glycerol yielded survival rates of 53.8% for D-shaped larvae and 37.5% for umbo larvae. The surviving D-shaped larvae showed active rotary motion and perfect membrane integrity and cytoplasmic normality, and the vigorous movement of veliger cilia was observed inside the closed shells. The breakdown of tissue occurred in the abnormal larvae, and the isolated cell often run out of shells.

• Proceedings of the Korean Society of Embryo Transfer Conference
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• 2002.11a
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• pp.104-104
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• 2002

This study was performed to find out the optimal larval stage among trochophore, D-shaped and umbo stage larvae and the desirable protective additive such as fructose, glucose, sucrose and trehalose with cryoprotectant for cryopreservation of surf clam, Spisula sachalinensis larvae. Dimethyl sulfoxide and ethylene glycol were used as cryoprotectant and each cryoprotectant was made to 2.0 M with previous protective additives. The larvae were immersed in the preparations waited for 15 minutes to reach equilibration, and then frozen in a program freezer (-35$^{\circ}C$) and liquid nitrogen (-196$^{\circ}C$). The freezing rate of 1.0$^{\circ}C$ /min. was used for cryopreservation of trochophores before seeding temperature (-12$^{\circ}C$). The survival rate of frozen-thawed larvae increased as larval developing and that of umbo stage larvae was the highest as 96.1 ${\pm}$ 1.0%. The presence of lower concentration of disaccharides as sucrose or trehalose significantly enhanced survival rate when mixed with cryoprotectants (P<0.05). The results of our study indicate that desirable developmental stages of larvae and protective additive for cryopreservation are the umbo stage larvae and 0.2 M sucrose, respectively.

• Proceedings of the Korean Society of Embryo Transfer Conference
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• 2003.10a
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• pp.72-72
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• 2003

• Korean Journal of Fisheries and Aquatic Sciences
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• v.18 no.5
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• pp.471-476
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• 1985

In order to set up a predictive model for effective spat collection of Pacific oyster, Crassostrea gigas, the survival rate and time required at each developmental stage of drifting larvae were surveyed during the period from June 13 to October 16 in 1984 at the main oyster farming area, Hansan Bay, the southern part of Korea. The advent of D-shape larvae ca. $72{\times}61{\mu}m$ long had two peaks in that area : July 16 and September 20. Umbo-shape larvae ca. $189{\times}217{\mu}m$ and full grown larvae co. $303{\times}308{\mu}m$ long also showed two peaks: July 21 and September 25 for the former, and July 28 and September 30 for the latter. About five days were required for D-shape larvae to develop to umbo-shape larvae. The instantaneous death rate was 0.24 and the daily survival rate 0.78 with a mean survival rate of $29.38\%$, at this intermorphological stage. The turnover time of umbo to full grown larvae varied from five to seven days with a instantaneous death rate of 0.15, daily survival rate of 0.86 and mean survival rate of $38.54\%$. Ten to twelve days were required for each group of the D-shape larvae to reach a full grown stage, and their mean survival rate was $11.32\%$ during this developmental period.

• Proceedings of the Korean Society of Embryo Transfer Conference
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• 2002.11a
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• pp.103-103
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• 2002

This study was conducted to investigate cryopreservation of pearl oyster, Pinctada fucata martensii larvae. Four cooling rates (-0.25, -0.5, -0.75 and -1.0$^{\circ}C$/min.) were used to examine a proper cooling rate during cryopreservation of trochophores before seeding temperature (-12$^{\circ}C$). Seven developmental stages (early and late trochophores, early and late D-shaped larvae and early, middle and late umbo stage larvae) and different sugars (fructose, glucose and sucrose) were used to investigate optimal larval stage and effective sugar in cryopreservation of larvae. The survival rates of frozen-thawed trochophores increased at cooling rate of -1.0$^{\circ}C$/min. As larval developing, survival rate of frozen-thawed larvae increased, except umbo stage larvae, and especially late D-shaped larvae highly survived as 91%. Addition of sugar revealed positive effect on cryopreservation in this experiment and 0.2 M glucose and sucrose mixed with 2.0 M dimethyl sulfoxide significantly enhanced survival rate of larvae (P<0.05). The results of our study indicate that desirable cooling rate, developmental stages of larvae and effective sugar far cryopreservation of pearl oyster, P. fucata martensii larvae are -1$^{\circ}C$/min, late D-shaped larvae and 0.2 M glucose and sucrose, respectively.

• Development and Reproduction
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• v.17 no.4
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• pp.337-343
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• 2013

Post-thawed larval rearing in Pacific oyster Crassostrea gigas was performed to investigate the survival rate with time course in three kinds of larvae cryopreserved. The highest survival rate and larval activity index (LAI) of post-thawed larvae were obtained from the permeation in 0.2 M sucrose and 2.0 M ethylene glycol (EG) at $-1^{\circ}C/min$ in freezing speed showing the survival rates just after thawing of 63.8% in trochophore, 84.1% in D-shaped veliger and 56.3% in early umbo veliger. In post-thawed larval rearing with food supply, the larvae lasted their lives until 24 hours in trochophore, 75 hours in D-shaped veliger and 57 hours in early umbo veliger. The results suggested that each larval stage post-thawed revealed no more further development to subsequent respective stage.

• The Korean Journal of Malacology
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• v.15 no.2
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• pp.115-119
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• 1999

We investigated the processes of egg and larval developments for aquaculture technique development of seedling production fo the flat oyster, Ostrea denselamellosa. Teo flat oyster of larviparous type was different from the pacific oyser (ovivarous type) because their larvae (trochophore and prodissoconch larvae) in the gill released into the seawater. The process of egg development was observed by artificial fertilization at $25^{\circ}C$, using a dissecting method. The sizes of Unfertilized eggs ranged from 80 to 90 $\mu\textrm{m}$ and fertilized eggs with globule-shape was 90-100 $\mu\textrm{m}$. The Polar body appeared after fertilization and egg cleavage began within 1 hour, reaching the blastula stage after 10 hours. The trochophore in the gill appeared 2-3 days after fertilization and grew to the prodissoconch larvae (130 140 $\mu\textrm{m}$) having a complete shell after 1-2 days. The shell of prodissoconch larvae grew to 205 220 $\mu\textrm{m}$ after 10 hours, and then they became umbo stage larvae showing oval in shape. The velum of umbo stage larvae was degenerated about 17-20 days after fertilization and grew into a pediveliger with a developed foot, at this time, the shell length size was 320 360 $\mu\textrm{m}$.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.21 no.1
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• pp.35-41
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• 1988

In order to set up a predictive model for effective spat collection of sea mussel Mytilus edulis , the survival rate and time required at each developmental stage of drifting larvae were surveyed during the period from March 14 to July 20 in 1987 at the Naesan Ri, Jinhae Bay, the southern part of Korea. The advent of D - shape larvae ca. $120\times90um$ um long had three peaks in that area: April 15, May 13 and June 7. Umbo shape larvae ca.$188\times162{\mu}m$ and full grown larvae ca. $289\times280{\mu}m$ long also showed three peaks: April 27. May 24 and June 20 for the former, and May 10, June 5 and June 30 for the latter. Eleven to thirteen days were required for D - shape larvae to develop to umbo - shape larvae. The instantaneous death rate was 0.1300 and the daily survival rate 0.8781 at this intermorphological stage. The turnover time of umbo to full grown larvae varied from ten to thirteen days with a instantaneous death rate of 0.1520, daily survival rate of 0.8590, and mean survival rate of $16.89\%$. Twenty - three to twenty - five days were required for each group of the D - shape larvae to reach a full grown stage, and their mean survival rate was $3.55\%$ during this developmental period.

• Development and Reproduction
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• v.5 no.1
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• pp.35-38
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• 2001

This study was peformed to find out the optimum larval stage among trochophore, early Dshaped larva, late D-shaped larva, early umbo and late umbo stages for cryopreservation of surf clam, Spisula sachalinensis larvae. Dimethyl sulfoxide (DMSO)and ethylene glycol were used as cryoprotectant, The larvae were immersed to cryoprotectants for 10 minutes and thereafter, cryopreserved in liquid nitrogen. Survival rates of trochophores frozen-thawed in 2.0 M dimethyl sulfoxide and 2.0 M ethylene glycol were the highest as 97.4% and 85.0%, respectively and post-thaw survival rates were decreased with the larval growth.