KR100571997B1 - Method of collecting and culturing larvae of korean soft corals - Google Patents

Abstract

The present invention provides a larval harvesting method and culture method for mass growth of Korean soft corals.

Algae larvae collection method of the present invention,

1) selecting a mature entity of the operational call; 2) extracting the selected operational call; 3) culturing the collected algae to generate larvae; And 4) harvesting the larvae.

Coral larvae cultivation method of the present invention is characterized by culturing while supplying water, oxygen, calcium and food into the culture water.

Conditions for developing algae larvae into polyps and colonies by obtaining the larvae generated by oil breeding of the algae through the algae larval harvesting method of the present invention, and culturing the obtained larvae by the algae cultivation method of the algae algae of the present invention. By optimizing this, it is possible to mass-produce Korean coral resources, which can be used directly as a means of securing species for conservation of species and restoration of ecosystems, as well as new material research and genetic research. This brings about an increasing effect.

Description

Method for collecting larvae and cultivating larvae of Korean coral reefs {Method of collecting and culturing larvae of Korean soft corals}

1 shows the frequency of monthly mature seminal vesicles of large resin cockscomb,

Figure 2 shows the frequency of monthly mature oocytes of large resin cockscomb,

Figure 3a is a photograph showing the development of the large resin cockscomb, Figure 3b is a change in the frequency of individuals in the P stage, OP stage, WP stage in the initial occurrence of large resin cockscomb,

Figure 4a is a photograph showing the development of the chestnut mandrami, Figure 4b shows the change in the frequency of individuals in the P stage, OP stage, WP stage in the early occurrence of chestnut mandrami,

Figure 5 shows the survival rate of the larvae according to the culture number in the algae larva culture,

Figure 6 shows the survival rate of the larvae according to the conditions affecting the development in the algae larva culture.

The present invention relates to a larval harvesting method and larval culture method for mass propagation of Korean algae.

Coral reptiles are called "flowers of the sea" and have been commercialized for jewelry and jewelry for a long time, but their value has recently been reevaluated as the anti-cancer and antibacterial effects of coral extracts have been reported. Corals are designated by the Convention on International Trade in Endangered Species of Wild Fauna and Flora (CITES). In Korea, 15 species of coral reptiles were designated as protected species among 'endangered and protected wild animals and plants in Korea' in 1998.In 2004, 'Yonsan Lake Community' and 'Seaweed' were designated by the Ministry of Cultural Heritage. It was designated as a natural monument. Coral reef formation in Earth's history has created an environment where living things can live on the earth, and coral habitats now form fisheries and form an ecologically important marine ecosystem where various creatures can live together.

In the temperate seas of Korea, Seogwipo Munseom, Forest Island and Beomseom Island in the sea area of Jeju Island form a unique marine ecosystem in which various marine life is distributed under the influence of Kuroshio turbulence. The region is well-developed in subtidal rock formations, and is home to a variety of adherents, especially corals and marine species, and is considered to be the region with the highest species diversity in Korea. The region's spectacular soft coral community is a unique ecosystem not found in tropical or subtropical regions, which was previously reported to UNEP's International Coral Reef Initiative in 2000 and is also protected by the World Marine Conservation Area Commission. We are planning to declare and protect the Marine Protected Area (MPA).

However, in recent years, valuable coral lakes have been rapidly destroyed by industrial activities such as reckless harvesting, fishing, submarine tourism, and breakwater construction. In addition, coral reptiles have been depleted of valuable coral resources for a long time as various jewels and decorations, and endangered as they are used for marine tourism or aquarium.

In Florida, Hawaii, and Guam of the United States, large-scale coral farms are being developed and produced as a preservation business, ornamental, and commercial resources, but large-scale breeding of coral reefs is being carried out. have.

However, no attempt has been made to multiply corals in Korea, and only a few aquarium shops have imported some foreign coral species to decorate their tanks. In addition, the Korea Maritime Research Institute's research project for the development of some ornamental species, "Basic research for the development of marine marine coronary organisms in Korea," is not a food resource, but is an early stage of ecological research and development of aquaculture and farming for other purposes. It is just fulfilling.

On the other hand, Korean Patent Laid-Open No. 10-2005-3508 (published on Jan. 12, 2005) relates to a method of mass propagation by asexual reproduction of Korean coral reptiles, and discloses propagation by asexual reproduction of aquatic corals. Proliferation by asexual reproduction according to this method, it is easy to obtain proliferative individuals from the mother, but these are all clones, which can reduce the genetic pool in the habitat, leading to a decrease in genetic diversity. In addition, the asexual reproduction method has a disadvantage in that the number of proliferative individuals that can be obtained from the parental 1 individual is limited.

Therefore, it is required to develop a method for mass proliferation of coral algae by sexual reproduction, and for this purpose, a method of mass propagation of mature algae can be determined by studying the reproductive patterns such as reproductive cycle and reproductive method of the breeding target species. .

Accordingly, the present inventors have found an optimal method for proliferating and culturing Korean algae in large quantities, thereby completing the present invention.

The present invention is to provide a larval sampling method and culture method for the mass growth of Korean algae.

 The present invention provides a method for collecting larvae for mass growth of Korean soft corals.

Algae larvae collection method of the present invention,

1) selecting a mature entity of the operational call; 2) extracting the selected operational call; 3) culturing the collected algae to generate larvae; And 4) harvesting the larvae.

The arithmetic corals may belong to all arithmetic corals, and may be one selected from a large resin mandrami ( Dendronephthya gigantea ), a chestnut mandrami ( Dendronephthya castanea ), and a purple resin mandrami ( Dendronephthya putteri ).

The algae larvae collection method of the present invention is to collect the larvae generated by the meteor breeding of the algae in the form of gonochoric or hermaphroditic.

In the first step of the algae larvae collection method of the present invention, the male and female maturation of the algae is selected.

In the case of large resin cockscomb, it is preferable to select males, and their sexual reproduction period is summer to early autumn and mature individuals are selected from June to October.

Oocytes mature in June-October and seminal vesicles in June-September. During this period, the main sexual reproductive period is July to September, when mature oocytes are the most frequent and larvae occur in female colonies through sexual reproduction in July to October. ~ October is the most desirable

Mature oocytes of the large resin mandrami are 300-500 μm in size and are orange in color. Oocytes are enclosed in vesicles in a circular shape, the inside of the gonad is filled with egg yolk, and the nucleus is biased to one side.

Mature seminal vesicles of large resin cockscomb are 250 ~ 375 ㎛ in size and are characterized by ivory color. The seminal vesicles are irregular in shape compared to oocytes. When immature, the immature cells completely fill the seminal vesicles, but as they mature, the seminal vesicles are arranged in the middle of the seminal vesicles and become hollow in the center.

In the case of the night resin cockscomb, it also shows a sexual reproduction similar to the big resin cockscomb, and the main reproduction time is July to September.

In the second step of the arithmetic coral mass breeding method of the present invention, the selected arithmetic coral is collected, and it is preferable to cut some branches of the mature aquatic colony selected in the first stage to a size of 8 to 10 cm.

Use sharp scissors or a knife as a tool for cutting at the time of collection.

The branch of cut algae is wrapped in vinyl using natural seawater and 100% oxygen, and the moved branch is temperature-adjusted at 23 ~ 26 ℃ for about 30 minutes.

The third step of the mass production method of operational coral in the present invention is a step of culturing the mature operational algae collected in the second step to generate larvae,

First, the cultured algae collected in the second step is first acclimated for about 24 hours in an oil-water culture tank containing natural seawater, and then cultured, and can be cultured in both natural seawater and artificial seawater.

When culturing algae in natural seawater, perform the culturing step in natural seawater immediately after acclimatization, and in the case of culturing algae algae in artificial seawater, go through the second step of acclimatization in mixed culture water of natural seawater and artificial seawater. To do that.

When culturing in natural sea water in the third step of the present invention, a) step of acclimatizing the harvested algae for 24 hours in a closed culture tank containing natural sea water; And b) culturing the acclimatized corals by replacing natural seawater at intervals of 12 hours in a closed culture tank using natural seawater to generate larvae.

When culturing in artificial seawater in the third step of the present invention, a) the step of firstly acclimatizing the harvested algae for 24 hours in a closed culture tank containing natural seawater; b) secondary acclimation of the first adapted corals for 24 hours in a closed culture tank containing culture water mixed with natural and artificial sea water, preferably in a 1: 1 ratio; And c) culturing the second adapted corals while exchanging artificial seawater at intervals of 12 hours in a closed culture tank using 100% artificial seawater to generate larvae.

The system of the culture tank is preferably a closed culture tank having a capacity of 5-10 L, and since a proper water flow is essential for culturing the algae, it is preferable to install a large-capacity oxygen feeder for water flow and oxygen supply.

In addition, a device that filters the culture tank and a skimmer, a protein remover, should not be used.

Since larvae are affected by factors such as sea temperature and ammonia, they should be adjusted to the same conditions as the habitats below (Bassim et Sammarco, 2003).

[Habitat quality conditions at the time of reproduction]

Ammonia (NH ₃ , NH ₄ ): 0.023 ~ 0.038 mg / l

Nitrite (NO ₂ ): 0.001-0.009 mg / l

-Nitrate (NO ₃ ): Always 0

-Temperature: Maintain 23 ~ 26 ℃

-Hydrogen ion concentration (pH): maintained at 8.1 ~ 8.3

Salinity: 32 ~ 33 ‰

Calcium: 350 ~ 400 ppm

In addition, in the present invention, as a system of a culture tank for generating larvae, a tank different from a general culture tank system should be used. First, the culture tank is preferably a closed culture tank having a capacity of 5 to 10 L. Since proper water flow is essential for cultivation, it is preferable to install a large-capacity oxygen supply for water flow and oxygen supply.

In addition, a device that filters the culture tank and a skimmer, a protein remover, should not be used.

In the present invention, the operational corals include a 'broadcasting mode' for releasing a gamete and an 'internal brooding mode' for releasing a planulae larva and an 'internal brooding mode'. external surface brooding mode '(Benayahu, Hydrobiologia, 216/217, 125-130, 1991).

In the case of large resin cockscomb, larvae are generated in female colonies through sexual reproduction, and larvae are released in the form of 'internal brooder', which is wrapped in mucus and released.

Chestnut cockscomb also exhibits a sexual reproduction similar to the large resin cockscomb and releases larvae in the form of 'incubator'.

In the fourth step of the algae larvae collection method of the present invention, the larvae generated and released in the step 3 is collected, the growth larvae by filtering the culture water using a plankton network having a grid size of 100 ~ 300 ㎛ It is desirable to obtain.

The larvae released from the mothers collected in step 4 of the present invention are orange in size 500-1200 ㎛ and rice grains, the larvae released clearly show the bicotyledonous tissue, and has many cilia on the body surface Seems.

The larvae of the algae harvested in this way can be grown and developed into polyps by culturing through the larval culture method described below.

The present invention provides a larval culture method for mass production of Korean algae.

The present invention provides a soft coral larvae cultured larvae cultured while supplying water, oxygen, calcium and food into the larvae.

In the algae larva cultivation method of the present invention, the larvae are generated by the sexual reproduction of the algae, the algae are all possible to belong to the algae, large resin mandra ( Dendronephthya gigantea ), chestnut mandra ( Dendronephthya castanea ) and Dendronephthya putteri are preferred. Moreover, it is preferable to collect | collect from the sea near Seogwipo, and it collects in Munseom and Beomseom.

For the larval culture, it is desirable to give the culture conditions in accordance with the habitat condition of Korean algae inhabited, and to find the optimum conditions for growth according to the species through various condition experiments during the process.

In the larval culture method of the present invention, the culture water may be used both artificial seawater or natural seawater.

Cultured water affects the growth rate and survival rate of larvae. For high larval viability, natural seawater is preferable, and natural seawater filtered through a 0.45 μm filter is most preferable. In addition, it is preferable to change culture water every 12 hours.

Incubation temperature of the larvae is preferably 22 ~ 24 ℃ considering the seawater temperature of the reproduction time.

In addition, larvae can be cultured in both closed and oil tanks, but since larvae have a floating planktonic larva that floats underwater until they are attached to a substrate and metamorphose, It is desirable to prevent larvae from escaping.

The cultured water is characterized by culturing while supplying the appropriate water flow and oxygen, it is preferable that a large-capacity oxygen supplier is installed for this purpose.

In addition, in the larval culture method of the present invention, it is preferable for normal development of the larvae that the food and calcium administration are properly controlled according to the larval stage of development.

In the culture method of the present invention, the feeding of the larvae (feeding) is characterized by feeding on phytoplankton of 1 ~ 2 ㎛, it is preferable to administer the feed after the white primordial polyp (WP) step in culture.

In the culture method of the present invention, it is preferable to contain and maintain the calcium concentration in the culture water containing 500 mg / liter. Calcium is essential for the growth of the algae in the form of polyps in the flanula larvae, and is most preferably administered to the culture water to maintain a 500 mg / liter concentration throughout the culture.

In the larval culture of algae, the larvae exhibit a certain level of activity until the larvae adhere to the substrate and metamorphose.

In the first stage, larvae move freely up, down, left and right in the culture water.

The second step is to swim just below the surface.

In the third step, it descends to the bottom to find a suitable substrate for attachment.

After the planulae (P) larvae adhere to the substrate, they begin metamorphosis and grow through the orange primordial polyp (OP) to the first white primordial polyp (WP).

The OP stage polyps have tentacle grooves for the development of idol-free tentacles and developing diaphragms, diaphragms, gastric ducts, and the like.

Polyps in the WP stage develop into complete initial polyps with the formation of tentacles, pinnule, septum, septum, gastric ganglia, and bone fragments.

After the growth of a complete polyp as an individual, the second polyp is produced by the germination method, which is asexual reproductive method, and continuously undergoes the developmental stage of asexual growth by colonization.

By obtaining the larvae generated by the breeding of algae in the algae larvae harvesting method of the present invention, and by developing the algae larvae into polyps and colonies through the optimized larval culture method of the present invention, a large amount of coral corals in Korea It can be used directly as a means of securing species for conservation of species and restoration of ecosystems, as well as for research of new materials and genetics, and increases economic feasibility through domestic and export of ornamental marine aquarium projects.

Hereinafter, preferred examples are provided to aid in understanding the present invention. However, the following examples are merely provided to more easily understand the present invention, and the contents of the present invention are not limited by the examples.

EXAMPLES Larval Sampling and Larval Culture of Korean Algae

1) Investigation of reproductive cycle and timing of computational corals

In order to select and harvest mature individuals of the algae for mass propagation by the sexual propagation of the algae of the present invention, a study was conducted on the reproductive cycle and the reproductive time of the algae.

About 100 gonads were extracted each month from the colony of large resin cockscomb which is a proliferation target using scuba in Munseom and Beomseom near Seogwipo and examined the size, shape and color of gonads. This experiment was conducted from February 2003 to January 2004.

Mature gonads were distinguished by size, internal morphology, and color change, and the unit of size was expressed in μm.

The results are shown in FIGS. 1 and 2.

As shown in FIG. 1, the large resin cockscomb has a high frequency of mature seminal vesicles in June and September, respectively, and as shown in FIG. 2, mature oocytes appear to have a high frequency in June and October. It was investigated.

In addition, females had the mature gonads at the lowest frequency in June, whereas males had the highest frequency of mature gonads in June, indicating that males reached sexual maturity before females. Mature oocytes are most commonly observed in July and September, indicating that this is the main reproductive period.

In addition, mature oocytes were 300-500 ㎛ in size and orange in color. Mature seminal vesicles had an ivory color of 250-375 ㎛ in size. Oocytes are round and wrapped in vesicles. The inside of the gonad is filled with egg yolk and the nucleus is biased to one side. The seminal vesicles have an irregular shape compared to oocytes. When immature, the garden cells fill the seminal vesicles, but as they mature, the seminal cells are arranged in the middle of the seminal vesicles, making the central region appear empty.

In order to identify sexual reproduction, histological and field surveys were conducted in parallel with the reproductive period estimated by the above-described gonad developmental status survey.

As a result, the large resin cockscomb was found to be a breeder in the body that releases larvae in mucus between July and October.

In addition, the protected species Bam Suzymandrami confirmed that the reproductive period is July ~ September in the same way.

2) Collecting larvae for the mass breeding of algae in the present invention

During the sexual reproduction of the large resin mandrami and chestnut mandrami investigated above, some branches of female colonies having larvae in Munseom and Beomseom near Seogwipo were cut and cut to 9 cm in size using sharp scissors.

The cut eggplant was transported in vinyl using natural seawater and 100% oxygen, and the transported eggplant was subjected to temperature adaptation for about 30 minutes in a packaged state, and then in a flowable natural seawater culture tank for about 24 hours. Tea was acclimated.

After acclimatization, both natural and artificial seawater can be cultivated, and when larvae are collected from artificial seawater, a second step of acclimation is required.

When cultivating the collected algae in artificial seawater, transfer the first acclimated branches to a 7-liter closed culture tank that supplies oxygen to the culture water mixed with natural seawater and artificial seawater 1: 1. After the passage, the cells were cultured in a closed tank using 100% artificial seawater. After transferring to a closed culture tank, the culture water was changed twice a day at 12 hour intervals, and the growth larvae were collected by filtering the culture water using a plankton network having a grid size of 100 μm.

The larvae emitted from the algae branch were orange, 500-1200 μm in size, and shaped like rice grains (see FIGS. 3A and 4A). The released larvae clearly showed bilobal tissue and had a large number of cilia on the surface of the body.

Larval outbreaks are affected by factors such as sea temperature and ammonia, so they should be matched to the conditions of habitats where possible (Bassim et Sammarco, 2003).

3) Condition optimization experiment for culturing algae larvae of the present invention

In the present invention, in order to increase the success rate of mass production of the algae through sexual reproduction, the optimum conditions for larval culture of the algae was examined.

Large resin cockscomb and chestnut cockscomb larvae prepared in Example 1 were used, and the experimental conditions were tested whether artificial seawater / natural seawater and food / calcium affected the survival rate and growth rate of larvae and polyps during the initial development. Was performed.

At this time, the early stages of larvae were divided into three stages to facilitate the development of each condition.

The first is the Planulae larval stage, denoted by P in the results table, and the developmental status of this phase includes the movement of larvae until they adhere to the substrate and transform into polyp stages.

The second is the initial polyp stage, which is marked as OP (orange primordial polyp) in the result table. This step is until the larva forms a tentacle with an idol during the transformation into polyps. The polyps are pale yellow at this time, and tentacle grooves are formed to create tentacles.

The third is the late polyp stage, which is marked as white primordial polyp (WP) in the result table.

Figures 3a and 3b shows the change in the frequency of the individuals in the P, OP, WP phase of the development pattern and early developmental stage of large resin cockscomb,

4A and 4B show changes in the frequency of individuals in the P, OP, and WP stages of the developmental pattern and early developmental stages of chestnut cockscomb.

3-1) Comparison of Artificial Seawater / Natural Seawater Effects During Early Development

In order to increase the success rate in the early development process, the following experiments were conducted to determine the degree of influence on the incidence rate of artificial seawater and natural seawater.

The larvae were cultured in artificial seawater culture water and natural seawater filtered with a 0.45 μm filter in sterile 6-well, respectively, in a 24 ° C. growth chamber.

By the number of cultures, the artificial seawater developed from the P stage to the OP and WP stages within two days, and in the natural seawater, it was developed into polyps in the WP stage within two days of cultivation. Turned out to be absent.

However, the survival rate according to the culture conditions was found to be different depending on the stage of development.

The survival rate according to the culture water of the algae larva of FIG. 5 is higher in natural seawater in the early stage of development, but higher in artificial seawater in the middle stage, and higher in natural seawater in the late stage, and overall in natural seawater. Survival was high.

3-2) Comparative study of food / calcium effects during early development

According to the results of 3-1), it was estimated that various requirements necessary for development were more important in the late stage of development, and the food and calcium which greatly influenced the survival and growth of algae larvae were investigated.

Phytoplankton of 1 ~ 2 ㎛ size was used, and calcium was maintained at 500 mg / l, which is about 100 mg / l higher than the concentration of natural seawater.

The experiment was carried out in the experimental group treated with only one condition of feeding or calcium conditions in the natural seawater filtered with 0.45 ㎛ filter, and the experimental group treated with both conditions of feeding and calcium.

In the results of Figure 6, it was found that the food and calcium increases the growth rate in the development process. The survival rate was 25% after 11 days of culture in the feeding conditions and 75% and 38.8% in the calcium and feeding / calcium conditions, respectively, indicating that the calcium condition had the greatest effect on the survival rate.

In addition, on the 14th day of culture, the frequency of WP was more than two times higher in the food / calcium condition than in the other conditions.

As described above, according to the larval harvesting method of the present invention, the individual having a mature gonad during the sexual reproduction period of the algae is collected, and the algae larvae are conditioned according to the larval cultivation method of the present invention. It is possible to develop and mass proliferation was confirmed.

Conditions for developing algae larvae into polyps and colonies by obtaining the larvae generated by oil breeding of the algae through the algae larval harvesting method of the present invention, and culturing the obtained larvae by the algae cultivation method of the algae algae of the present invention. By optimizing this, it is possible to mass-produce Korean coral resources, which can be used directly as a means of securing species for conservation of species and restoration of ecosystems, as well as new material research and genetic research. This brings about an increasing effect.

Claims (10)

Soft coral larvae The larvae larvae cultured by supplying water, oxygen, calcium and food into the culture water. The method of claim 1, wherein the algae is one selected from a large resin mandrami ( Dendronephthya gigantea ), chestnut mandrami ( Dendronephthya castanea ), purple resin mandrami ( Dendronephthya putteri ). The larval culture method of claim 1, wherein the culture water uses artificial sea water or natural sea water. 4. The larval culture method according to claim 3, wherein the cultured water uses natural seawater filtered with a 0.45 µm filter. The larval culture method according to claim 1, wherein the culture water is exchanged every 12 hours. The method of claim 1, wherein the larvae feed larvae cultured characterized in that feeding on 1 to 2 ㎛ phytoplankton. The larval culture method of claim 1, wherein the larval food is administered after a white primordial polyp (WP) step in culture. The method of claim 1, wherein the calcium is contained in the culture water at a concentration of 500 mg / liter larvae culture method, characterized in that to maintain it. The method of claim 1, wherein the calcium is cultured larvae characterized in that the administration to the culture water throughout the culture. 10. The method according to any one of claims 1 to 9, wherein the larvae are harvested according to the following steps; 1) selecting a mature entity of the operational call; 2) extracting the selected operational call; 3) culturing the collected algae to generate larvae; And 4) harvesting the larvae.

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