• Journal of Animal Reproduction and Biotechnology
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• v.34 no.4
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• pp.280-291
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• 2019

Potential utility of 14 candidate housekeeping genes as normalization reference for RT-qPCR analysis with developmental samples (fertilized eggs to late veliger larvae) in Pacific abalone Haliotis discus hannai was evaluated using four different statistical algorithms (geNorm, NormFinder, BestKeeper and comparative ΔCT method). Different algorithms identified different genes as the best candidates, and geometric mean-based final ranking from the most to the least stable expression was as follow: RPL5, RPL4, RPS18, RPL8, RPL7, UBE2, RPL7A, GAPDH, RPL36, PPIB, EF1A, ACTB and B-TU. The findings were further validated via relative quantification of metallothionein (MT) transcripts using the stable and unstable reference genes, and expression levels of MT were greatly influenced according to the choice of reference genes. In overall, our data suggest that RPL5 and RPS18, either singly or in combination, are appropriate for normalizing gene expression in developmental samples of this abalone species, whereas ACTB, B-TU and EF1A are less stable and not recommended. In addition, our findings propose that standard deviations in geometric ranking as well as geometric mean itself should also be taken into account for the final selection of reference gene(s). This study could be a useful basis to facilitate the generation of accurate and reliable RT-qPCR data with developmental samples in this abalone species.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.9 no.1
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• pp.43-55
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• 1976

Fertilization and early development of turbo cornutus was studied based on the samples which were collected in Yeosu area. Particular emphasis was paid on induction of artificial spawing, fertilization rate, preembryonic development, the growth of the early larva and larval survival to various salinity. Among the various methods for induction of artificial spawning which have been tested for the present study, drying by exposure to air is the. most efficient, and percentage fertilization rate was $83.8-96.4\%$. The diameter of fertilized eggs was $0.182{\pm}0.0028mm$; and the diameter of egg membrane was $0.245{\pm}0.093mm$. Under the temperature range of $20.6-25.4^{\circ}C$ the larvae hatched out after 11:05-11:15 hours of fertilization. After 3.0-3.5 days of fertilization the planktonic larvae begand to settle, and the settlement terminated within 5 days. During the period of 150 days of early culturing the diameter growth of shell(M) and the diameter of shell aperture(A) was formulated as follows: $$1972\;M=0.33e^{0.02070D}$$ $$A=0.19e^{0.02282D}$$ $$1973\;M=0.32e^{0.02282D}$$ $$A=0.16e^{0.02596D}$$ During the same period of early culturing the relative growth of shell diameter and the diameter of shell aperture was formulated as follows : 1972 A=0.6478 S-0.1575 1973 A=0.5897 S-0.0515 After 11 days of larval hatching $0.02-0.18\%$ of planktonic larvae settled. After 150 days of settlement the survival rate of the early shells was $7.4-21.6\%$. Under the temperature range of $21.0-22.7^{\circ}C$ the optimum salinity range for the development of egg and the planktonic larvae was $30-35\%_{\circ}$.

• Journal of Aquaculture
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• v.15 no.2
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• pp.95-101
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• 2002

During the periods from lily to October, 2000 in Hongseong and lucy to October, 2001 in Taean in the west coast of Korea, the following environmental conditions prevailed : water temperature : 22.0~26.817, salinity 27.23 ~30.80%, dissolved oxygen 4.12 ~6.26 ml/l, pH 7.89 ~8.09, phosphate 0.39 ~0.65 $\mu m$ , inorganic nitrogen 5.05~9.26 $\mu m$, suspended solid 5.4~20.8 mg/l and chemical oxygen demand 1.12~1.87 mg/l. The B-shaped veliger larvae of the Ark shell occurred in maximum number at $25^{\circ}C$ prevailing from mid-August at Hongseong and Taean. Full grown larvae reached maximum abundance from late August. To identify the effectiveness of the substratum for spat collection, raschel net were tested to Larval settlement. The most effective depth to collect the larvae in natural environment was the collectors suspended at 7~8 m depth. At these depths, about 49 to 94 spats were found on the collector (40$\times$50 cm), The growth of shell height (Y) to shell length (X), and total weight (W) to shell length (L) could be formulated as follows respectively: Hongseong: SH = 0.7168 SL -0.6466 ( $r^2$ = 0.9839), TW = $0.0001SL^{3.1705}$ ($r^2$ = 0.9882) Taean: SH = 0.736 SL -0.8824 ($r^2$ : 0.9899), TW : 0.00005 $SL^{3.3731}$ ($r^2$ : 0.9899)

• The Korean Journal of Malacology
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• v.17 no.1
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• pp.45-55
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• 2001

Gametogenes, reproductive cycle, first sexual maturity(biological minimum size), sex ratio and larval development of the marsh clam Corbicula japonica were investigated monthly by histological observations. Samples were collected in brackish water of Gokgang stream, Kyungsangbuk-Do, Korea, from August 1997 to July 1998. Sexuality of Corbicula japonica is dioecious and the species are an oviparous clam. The gonads are irregularly arranged from the sub-region of mid-intestinal gland in visceral cavity to reticular connective tissue of foot. The ovary is composed of a number of ovarian sac which are branched arborescent. Oogonia actively proliferate along the germinal epithelium of ovarian sac, in which young oocytes are growing. The testis is composed of a number of testicular tubules, and the epithelium of the tubule has function of germinal epithelium, along which spermatogonia actively proliferate. A great number of undifferentiated mesenchymal tissue and eosinophilic granular cells are abundantly distributed between developing oocytes and spermatocytes in the early developmental stages. With the further development of the ovary and testis these tissue and cells gradually disappear. Then the undifferentiated mesenchymal tissue and eosinophilic granular cells are considered to be related to the growing of the oocytes and spermatocytes. The spawning period is from July to September, and the main spawning occur between July and August when seawater temperatures reach above 22$^{\circ}C$. The reproductive cycle of this species can be divided into five successive stages; early active (February to April), late active (May to July), ripe (June to September), partially spawned (July to September), degenerative (September to October) and resting stage (October to February). Percentages of first sexual maturity of female and male clams ranging in length from 10 mm to 12 mm are over 50% and 100% for clams over 16.0 mm in shell length. Fertilized eggs or Corbicula japonica were 80-90 ${\mu}{\textrm}{m}$ in diameter. In the early embryonic development of C. japonica, the appearance of polar body, trochophore and D-shaped veliger were observed around 40 min., 27 hours and 4 days after spawning, respectively, at a water temperature of 26.5-28.$0^{\circ}C$. The size of larvae of early umbo stage was about 185-210 ${\mu}{\textrm}{m}$ in shell length, 160-180 ${\mu}{\textrm}{m}$ in shell height around 7 days after fertilization. The correlation of relative growth between the culture day (D) and shell length (SL) was expressed by the following simple formula from D-shaped veliger to metamorphosing stage; SL = 13.300D + 209.36($r^2$= 0.9078).

• The Korean Journal of Malacology
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• v.21 no.1
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• pp.25-31
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• 2005

The embryos of marine bivalves have been commonly used in bioassays for the quality assessment of marine environments. Although several standard protocols for developmental bioassay with bivalves have been already proposed, there have been few trials for applying these protocols in environmental assessment, or for developing new protocol with Korean species. So, there is a strong need to establish the standard bioassay protocols using bivalves commonly found in Korean waters. Prior to developing a new protocol, it is essential to know the optimum conditions for the reliable bioassay procedures. Here, we established the purpose of this study to determine the optimum bioassay conditions for successful development of a common mussel, Mytilus galloprovincialis. The conditions considered as critical for developmental bioassay, and determined in this study were; (1) temperature, (2) salinity, and (3) initial density of embryo. The optimal temperature for developmental bioassay of M. galloprovincialis was determined as $15^{\circ}C$. At this temperature, the required time for the embryo to become veliger larva was 48 hr. The acceptable range of salinity for the embryotoxicity test using M. galloprivincialis was from 30 to 35 psu, which was narrower than that of the natural habitat of adult populations. The optimum density of embryo at the beginning of bioassay was 100 embryos/ml. Over this density, the proportion of normally developed larvae decreased significantly. The results obtained in this study will serve as a basis for preparation of the standard bioassay protocol using embryo of M. galloprovincialis.

• Development and Reproduction
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• v.6 no.1
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• pp.37-44
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• 2002

First sexual maturity, sex ratio, spawning frequency, deposition of the egg capsules and fecundity of the female Rapana venosa(Valenciennes) inhabited in the artificially closed slag deposit area, Gwangyang Bay were investigated by histologicai and visual observations for natural living resource management. The rate of individuals reaching the first sexual maturity was 51.6％ in females measuring 7.1~8.0 cm in shell height, and 100％ in those > 10.1 cm. The total number of egg capsules per individual and the mean number of eggs in an egg capsule were 192~382 and 500, respectively. However, the number of eggs per individual and sizes of egg capsules under lower salinity and deficient food conditions in the closed slag deposit area were smaller than those under the optimum salinity and sufficient food conditions in the open regions. Fecundities of the species were approximately from 96,000 to 191,000 eggs/individual with two to low broods(spawning frequencies) during the spawning season. The duration of development in egg capsules was 18~19 days at about 18~2$0^{\circ}C$. R. venosa is a species whose embryos hatch as veliger larvae, not juvenile snail. The sex ratio of female : male was not significantly different from 1 : 1($\chi$$^2$= 0.23, p>0.05).