Branchiostoma breed conservation method

Abstract

The invention provides a breed conservation method for branchiostoma belcheri. The method comprises the steps that S1, branchiostoma belcheri living bodies are collected from a natural sea area to serve as parent groups; s2, performing group breeding on the same group in a seawater environment paved with sea sand; s3, an algae nutrient solution is fed three times every day, water feeding is stopped for 2 hours after feeding, and the feeding interval is 7-9 hours; s4, emptying the cultivation seawater twice every day, wherein the drainage interval is 12 hours; s5, after it is found that the branchiostoma brevituba gets out of sea sand, swims in seawater or has special smell in air, the time of three times of feeding is adjusted, the interval of feeding every time is 6-7 hours, feeding is not carried out after 19: 00, the feeding amount every time is increased to 1.6-2 times that in the step S3, water does not enter after third-time feeding, and first-time feeding is carried out till the next day; s6, fertilized eggs in the seawater or larvae obtained by hatching the fertilized eggs are collected, and the larvae obtained by hatching the fertilized eggs are the first filial generation.

Description

Technical Field

[0001] The present invention belongs to the technical field of lancelet breeding, and relates to a method for preserving lancelet seeds. Background Art

[0002] Amphioxus is a chordate of the family Amphioxidae in the order Amphioxus. Amphioxus is of great academic significance because it is a transitional type from invertebrates to vertebrates, providing a valuable model for studying the origin and evolution of vertebrates. Due to overfishing and habitat destruction, the number of amphioxus once decreased.

[0003] It is of great significance to study and develop methods for the conservation of lancelets in order to provide guidance for the reproduction and protection of lancelets populations. Summary of the invention

[0004] The present invention provides a method for preserving the seeds of lancelets, which can effectively solve the above problems.

[0005] The present disclosure is implemented as follows:

[0006] In a first aspect, the present disclosure provides a method for preserving the seeds of lancelets, the method comprising:

[0007] S1, collect live lancelets from natural sea areas as parent populations, and select lancelets with fast movement and strong sand drilling ability;

[0008] S2: The same group of fish were cultured in a seawater environment with sea sand. The thickness of the sea sand was 5-10 cm. The water was continuously circulated and the water was seawater containing algae. The culture density was 6000-12000 fish / m 2 ;

[0009] S3, feeding algae nutrient solution three times a day, stopping water intake for 2 hours after feeding, and the interval between each feeding was 7-9 hours;

[0010] S4, drain the cultured seawater twice a day, with an interval of 12 hours between each drainage;

[0011] S5, if the lancelets were found to have emerged from the sea sand and were swimming in the sea water, or if there was a special smell in the air, they would not be fed with water after the third feeding of the day until the first feeding of the next day;

[0012] S6, after the third feeding in step S5, collecting fertilized eggs in the seawater, or larvae hatched from the fertilized eggs, the larvae hatched from the fertilized eggs are the first generation;

[0013] Algae-containing seawater: pH value is 8.3-8.5, ammonia nitrogen mass concentration is less than 0.05 mg / L, dissolved oxygen is higher than 5.5 mg / L, water temperature is 26-32°C, salinity is 19-29, and algae-containing seawater is obtained by filtering seawater from seawater ponds;

[0014] Algae nutrient solution: each feeding amount in step S3, ferric citrate 6-6.2×10 -5 ppm / L seawater, urea 1.2-1.3×10 -2 ppm / L seawater, potassium dihydrogen phosphate 6-6.2×10 -4 ppm / L seawater, 6000-10000 golden algae / mL seawater, or, replace no more than half of the golden algae with flat algae, and convert based on the volume ratio of golden algae to flat algae.

[0015] In a second aspect, the present disclosure provides a device for preserving the seeds of lancelets, which is used in the above method, and the device comprises:

[0016] A breeding facility, the bottom of which is paved with sea sand and the breeding facility is filled with sea water;

[0017] A drain outlet is provided at the bottom of the breeding mechanism, and a closing mechanism is installed on the drain outlet for closing the drain outlet;

[0018] A water inlet mechanism, the water inlet mechanism is arranged inside the breeding mechanism, the water inlet mechanism is provided with a water inlet, and the water inlet is arranged below the surface of the seawater in the breeding mechanism;

[0019] A water outlet mechanism, the water outlet mechanism is arranged inside the breeding mechanism, and a water outlet is arranged at the upper part of the water outlet mechanism;

[0020] The first filter screen is installed at the bottom of the breeding mechanism and is arranged at the outer side of the drain outlet.

[0021] The beneficial effects of the present disclosure are:

[0022] The present invention provides a method for preserving the seeds of lancelets, which can achieve the preservation of wild parents and can well simulate the natural growth environment of lancelets. The feeding adopts a method of combining continuous rough breeding and regular intensive breeding. The algae-sea water ensures that the lancelets can continuously obtain food, avoids excessive metabolic load caused by one-time intake of too much food through regular feeding, and fluctuations in metabolic levels. The algae nutrient solution realizes sufficient food supply, thereby improving the survival rate and growth rate of the lancelets.

[0023] Furthermore, when the lancelets are found to have signs of pre-mating behavior, suitable mating conditions and environment are provided by increasing the amount of feeding, feeding in advance, and stopping the seawater replacement cycle, thereby prompting them to mate, obtain fertilized eggs, and reproduce and cultivate the next generation.

[0024] Furthermore, by cross-pairing the first generation corresponding to different parents, the seeds are preserved and bred again to the second generation. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] In order to more clearly illustrate the technical solutions of the embodiments of the present disclosure, the drawings required for use in the embodiments will be briefly introduced below. It should be understood that the following drawings only show certain embodiments of the present disclosure and therefore should not be regarded as limiting the scope. For ordinary technicians in this field, other related drawings can be obtained based on these drawings without creative work.

[0026] Figure 1 It is a schematic diagram of the steps of the method for preserving the lancelet species provided in the embodiment of the present disclosure.

[0027] Figure 2 It is a first structural schematic diagram of the seed preservation device provided in an embodiment of the present disclosure.

[0028] Figure 3 It is a second structural schematic diagram of the seed preservation device provided in an embodiment of the present disclosure.

[0029] Figure 4 It is a structural schematic diagram of the storage mechanism of the device provided in an embodiment of the present disclosure.

[0030] Figure 5 It is a schematic diagram of using a collecting mechanism and a pumping mechanism to collect fertilized eggs in seawater in a breeding mechanism in the mating step in the method provided by an embodiment of the present disclosure, and using an aeration mechanism to aerate the seawater. DETAILED DESCRIPTION

[0031] In order to make the purpose, technical solutions and advantages of the embodiments of the present disclosure clearer, the technical solutions in the embodiments of the present disclosure will be clearly and completely described below in conjunction with the drawings in the embodiments of the present disclosure. Obviously, the described embodiments are part of the embodiments of the present disclosure, rather than all of the embodiments. Based on the embodiments in the present disclosure, all other embodiments obtained by ordinary technicians in this field without making creative work are within the scope of protection of the present disclosure. Therefore, the following detailed description of the embodiments of the present disclosure provided in the drawings is not intended to limit the scope of the present disclosure for protection, but merely represents the selected embodiments of the present disclosure. Based on the embodiments in the present disclosure, all other embodiments obtained by ordinary technicians in this field without making creative work are within the scope of protection of the present disclosure.

[0032] Example 1

[0033] like Figure 1 As shown, the embodiment of the present disclosure provides a method for preserving the seeds of lancelets, the method comprising:

[0034] S1, collect live lancelets from natural sea areas as parent groups, and select lancelets with fast movement and strong sand-drilling ability.

[0035] In some embodiments, in step S1, a lancelet with a body length of 35-55 mm, rapid movement, strong sand drilling ability, and full gonads is selected.

[0036] The collected wild lancelets are of different sizes, that is, the developmental degrees of the parent individuals are different. The collected parent individuals are group-raised without affecting seed preservation. The method can enable individuals with lower gonad maturity to reach a state of full gonads during the breeding process, and perform mating behavior to achieve the reproduction of the next generation.

[0037] Screening out strong individuals with fast movements and strong sand-drilling ability is to improve the success rate and efficiency of lancelet conservation. These individuals usually have strong vitality and reproductive ability, can better adapt to the artificial breeding environment, thereby improving the reproduction rate and survival rate of offspring.

[0038] The parent individuals have different degrees of gonadal development. Further screening out individuals with a body length of 35-55mm, rapid activity, strong sand-drilling ability, and full gonads for group breeding can shorten the seed preservation process, enable these individuals to pair and mate in a shorter period of time, and centrally collect fertilized eggs or young.

[0039] S2: The same group of fish were cultured in a seawater environment with sea sand. The thickness of the sea sand was 5-10 cm. The water was continuously circulated and the water was seawater containing algae. The culture density was 6000-12000 fish / m2 .

[0040] like Figure 2 and 3 As shown, an embodiment of the present disclosure provides a device for preserving the seeds of lancelets, which is used in the method.

[0041] The device comprises: a breeding mechanism. The parent is put into the breeding mechanism for breeding.

[0042] Specifically, the breeding facility is a PVC plastic barrel with a diameter of 1m and a height of 60cm. 7,000 lancelets are placed in the breeding facility.

[0043] The bottom of the breeding facility is paved with sea sand at a height of 5-10 cm to meet the habit of lancelets to drill into the sand. The height of the paved sea sand can bury the body of the lancelets.

[0044] The sea sand is taken from the sea area where lancelets live.

[0045] The particle size of sea sand is 0.5-1mm. Soak it in fresh water for 2 days and then soak it in fresh sea water twice.

[0046] The culture facility is filled with seawater, and the water level of the seawater is 50 cm.

[0047] The breeding mechanism is placed in the breeding room with natural light cycle and natural light intensity.

[0048] The seawater in the aquaculture facility is the algae-containing seawater.

[0049] The algae-containing seawater is natural algae-containing seawater. Using natural seawater can better simulate the natural growth environment of lancelets.

[0050] The algae-containing seawater is obtained by filtering the seawater in the seawater pond. The algae-containing seawater is taken from the seawater pond.

[0051] The seawater pond is an open water system dug in the seashore tidal flat. For example, the existing seawater pond can be used to breed shrimp ponds. The seawater in the seawater pond is filtered through a 300-mesh silk sieve to remove large particles such as sea sand and parasites before use.

[0052] The seawater in the seawater pond is obtained by pumping seawater that meets aquaculture standards from the natural environment into the pond. It contains natural algae, and the algae in the algae-containing seawater can grow naturally.

[0053] The stored algae-containing seawater is tested regularly to determine whether the content and proportion of algae therein meet the requirements for feeding lancelets and providing them with the nutrients needed for growth.

[0054] Seawater containing algae: pH value is 8.3-8.5, ammonia nitrogen mass concentration is lower than 0.05 mg / L, dissolved oxygen is higher than 5.5 mg / L, water temperature is 26-32℃, and salinity is 19-29.

[0055] In some embodiments, the algae-containing seawater has a content of 300-350 Platymonas / mL of seawater and a content of 2500-3500 Chrysophytes / mL of seawater.

[0056] Feeding lancelets with algae-containing seawater is a strategy for "rough farming" of lancelets.

[0057] Algae-containing seawater can continuously provide food for lancelets, allowing them to eat at any time, which is closer to their growth in a natural environment, rather than regular nutritional shocks, thereby improving their adaptability to the breeding environment and their survival rate.

[0058] Continuous feeding of algae-containing seawater can also reduce the amount of each feeding during scheduled feeding, reducing the impact of scheduled feeding on water quality, such as excessive turbidity of the water, which makes it difficult for lancelets to adapt to the changes in the breeding environment and unable to mate, or even die.

[0059] Studies have shown that water temperatures below 23°C can affect the mating behavior of some lancelet species.

[0060] like Figure 2 and 3 As shown, the device further comprises a water inlet mechanism. The algae-sea water is taken into the culture mechanism through the water inlet mechanism.

[0061] The flow rate of the incoming seawater should be such that the flow of seawater in the aquaculture facility can be observed.

[0062] In some embodiments, the flow rate of incoming seawater is 400-500 L / h.

[0063] The water inlet mechanism can be a PVC elbow with a diameter of 5 cm. Four of them are evenly arranged along the circumference of the breeding mechanism. The bending direction is along the same direction of the circumference, so that the incoming seawater forms a circulating water flow in the seawater in the breeding mechanism, so that the seawater in the breeding mechanism is evenly mixed.

[0064] Therefore, the number of water inlet mechanisms, the bending angle of the water inlet elbow, the position of the water inlet, and the water inlet flow rate are related to the height, cross-section and volume of the water body in the breeding mechanism.

[0065] By introducing seawater into the aquaculture facility to create a flow of seawater, the tidal fluctuations in the natural ocean environment can be simulated.

[0066] Moreover, by continuously replacing the seawater in the aquaculture facility through water inlet and outlet, a cycle is formed, which can increase the dissolved oxygen in the water and improve the water quality.

[0067] The incoming seawater is placed in the aquaculture room to a constant room temperature before entering the aquaculture facility.

[0068] Regularly test the seawater before it enters the water, such as the seawater in the water storage facility, to determine the indicators therein to avoid abnormalities in the seawater in the breeding facility that would cause the lancelets to be unable to survive.

[0069] In some embodiments, Figure 4 As shown, the device further comprises a water storage mechanism, and the water storage mechanism is connected to the water inlet mechanism.

[0070] The water storage mechanism and the breeding mechanism are both placed in the breeding room. Seawater is first taken into the water storage mechanism from the seawater pond, and then placed to a constant temperature before being used for the water intake mechanism.

[0071] Specifically, the water storage mechanism is a water tower, and the capacity of the water tower is 3-5 tons.

[0072] like Figure 2 and 3 As shown, the device further comprises a water outlet mechanism. The water outlet mechanism is arranged inside the breeding mechanism, and a water outlet is arranged on the upper part of the water outlet mechanism.

[0073] The water outlet mechanism can be a PVC straight pipe with a diameter of 5 cm. A water outlet is provided at the corresponding position of the water surface for drainage. Water is discharged while water is inletted to maintain the water level in the breeding mechanism.

[0074] S3, algae nutrient solution was fed three times a day, water intake was stopped for 2 hours after feeding, and the interval between each feeding was 7-9 hours.

[0075] By feeding algae nutrient solution to provide lancelets with sufficient nutrients, "intensive breeding" is achieved compared to the "rough breeding" of water containing algae and seawater to meet the growth and reproduction needs of lancelets.

[0076] The specific feeding times can be: 5:30, 14:30, and 22:30.

[0077] Algae nutrient solution: each feeding amount in step S3, ferric citrate 6-6.2×10 -5 ppm / L seawater, urea 1.2-1.3×10 -2 ppm / L seawater, potassium dihydrogen phosphate 6-6.2×10 -4 ppm / L seawater, 6000-10000 golden algae / mL seawater, or, replace no more than half of the golden algae with flat algae, and convert based on the volume ratio of golden algae to flat algae.

[0078] In some embodiments, 9 Chrysophytes are replaced with 1 Platymonas.

[0079] The main baits of the lancelets used are flat algae and golden algae.

[0080] In some embodiments, the phytoplankton is mainly composed of Qingdao phytoplankton. The golden algae is mainly composed of Zhanjiang phytoplankton.

[0081] Specifically, the method for preparing the algae nutrient solution comprises:

[0082] Three kinds of nutrient salt mother solutions were prepared respectively, with concentrations of: 0.3ppm ferric citrate, 60ppm urea, and 3ppm potassium dihydrogen phosphate.

[0083] First-level culture: Boil about 20 liters of seawater and cool it down, then add 10 mL of the prepared nutrient solution, pour in the cultured algae solution, let it expand, and carry out first-level inoculation culture. The first-level cultured flat algae tend to stick to the wall, and the golden algae tend to sink to the bottom. If you find the phenomenon of sticking to the wall or sinking to the bottom, you should actively shake the algae solution to make the algae evenly distributed. When shaking the algae solution, be careful not to use too much force to prevent the algae solution from splashing. Shake for about half a minute each time. A good algae solution is bright in color, lively, and suspended in a cloud-like state.

[0084] Secondary culture: In order to obtain more flat algae and golden algae, pour the algae liquid of the primary culture into a sterilized 50L plastic bucket for secondary culture. Add appropriate amount of filtered seawater and nutrients every day, and stir the water at least 4-5 times a day to ensure even distribution of the algae.

[0085] In some embodiments, the cultured algal liquid in the primary culture comprises a single algal species.

[0086] In some embodiments, the golden algae are not replaced with the platymonas in the algae nutrient solution.

[0087] In the process of lancelet conservation disclosed in the present invention, it was found that golden algae has the effect of accelerating the maturation of parents and improving the survival rate of seedlings.

[0088] Platycladus is rich in vitamins and fatty acids, which can provide the nutrients required for the growth of amphioxus. However, if only platycladus is fed for the conservation of amphioxus, it can be observed that female amphioxus do not carry eggs or carry a small number of eggs. If they do carry eggs, the overall appearance of the eggs shows that they are not mature enough.

[0089] The specific feeding amount should be based on the fact that the fish can basically consume all the food within the 2-hour water outage period.

[0090] In some embodiments, the feeding amount each time in step S3 is 3 mL of algae nutrient solution.

[0091] The purpose of stopping water supply is to prevent the algae nutrient solution fed from being lost with the water, so that the lancelets can fully eat during this period and obtain the nutrients needed for seed preservation.

[0092] In some embodiments, seawater within the aquaculture facility is aerated during water outages.

[0093] The device also includes an aeration mechanism, which is arranged inside the breeding mechanism.

[0094] Four aeration mechanisms may be evenly arranged along the circumference of the aquaculture mechanism so that the flow of seawater in the aquaculture mechanism can be observed.

[0095] The aeration rate, number of aeration mechanisms and aeration locations are also related to the height, cross-section and volume of the water body within the aquaculture facility.

[0096] Aeration can mix the seawater in the aquaculture facility evenly and maintain dissolved oxygen in the water, thus avoiding changes in the aquaculture environment, such as lack of circulation. At the same time, the addition of algae nutrient solution can cause a sudden change in local water quality, resulting in the lancelets not eating and being unable to obtain sufficient nutrition.

[0097] Whether it is through aeration or replacing the water inlet and outlet circulation to form water body fluctuations, it is also a simulation of the natural seawater environment to avoid changes in the aquaculture environment.

[0098] S4, drain the cultured seawater twice a day, with an interval of 12 hours between each drainage.

[0099] like Figure 2 and 3 As shown, a drain outlet is provided at the bottom of the breeding mechanism, and the diameter of the drain outlet is slightly larger than the PVC straight pipe for water discharge, for example, the diameter is 6 cm. A plug is installed on the drain outlet to close the drain outlet.

[0100] The PVC straight pipe used for water outlet can be directly installed on the plug and connected to the bottom of the plug, so that the plug can be pulled up through the PVC straight pipe to open the drain.

[0101] Furthermore, a first filter screen is arranged outside the drain outlet. The first filter screen is installed at the bottom of the breeding mechanism. The diameter of the first filter screen is slightly larger than the drain outlet, for example, the diameter is 7 cm.

[0102] The height of the first filter screen needs to be 1-2 cm higher than the height of the sea sand, so as to prevent the sea sand from being discharged from the drain port. The side of the first filter screen is provided with mesh holes so that the seawater in the sea sand can pass through.

[0103] The top of the first filter screen is open to provide a space for pulling out the plug.

[0104] The purpose of drainage is to simulate the rise and fall of tides in the amphioxus habitat. By draining the water, the sea sand is exposed to the surface, and then the incoming seawater will submerge the sea sand again.

[0105] The drainage time can be set between the first feeding and the second feeding, and between the second feeding and the third feeding, respectively, so that the seawater in the aquaculture structure can be filled again when the water intake is stopped next time.

[0106] Through drainage, the seawater in the breeding facility can be completely replaced, thus providing a better growth environment for lancelets.

[0107] Drainage simulates the self-purification effect of the marine environment, and twice a day is closer to the self-purification capacity of the marine environment.

[0108] Therefore, by adopting a mode of inlet and outlet water circulation and regular drainage, the upper limit of each feeding amount does not need to be controlled too precisely. Even if the feeding amount each time is slightly higher than the amount that lancelets can consume and there is residue in the seawater, there is no need to worry about the cumulative impact of multiple overfeeding on the concentration of nutrients such as algae in the seawater on the water quality.

[0109] In particular, based on this mode, during the concentrated mating period of the amphioxus in step S7, the feeding amount can be increased and the feeding time can be changed, while reducing the impact on the mating behavior of the amphioxus.

[0110] In some embodiments, before draining, the sea sand is stirred so that impurities in the sea sand float into the sea water and are discharged with the sea water.

[0111] After stirring, the lancelets in the sea sand will drill out of the sand layer, and then quickly drill back in after the stirring stops and the sand layer stabilizes. After drilling back, the water will be drained.

[0112] Impurities include food residues, metabolites of amphioxus, such as feces, and amphioxus carcasses.

[0113] Studies have shown that the cleanliness of the sand layer where lancelets live is crucial to their survival.

[0114] Impurities are removed to avoid affecting the sea sand environment where lancelets live, which would cause a decrease in the survival rate of lancelets.

[0115] Likewise, the removal of impurities with the water flow simulates the self-purification effect of the marine environment.

[0116] S5, when the lancelets were found to emerge from the sea sand and swim in the sea water, or when there was a special smell in the air, they would not take in water after the third feeding of the day until the first feeding of the next day.

[0117] Studies have found that the mating time of lancelets is between 20:00 and 24:00.

[0118] It was found during the disclosed process of lancelet conservation that this phenomenon generally occurs after dark, around 18:00 to 24:00 at night.

[0119] Therefore, if the third feeding has not been carried out after discovering this phenomenon, it can be done in advance.

[0120] The study found that when mating begins, the male fish jumps out of the water first and swims quickly, and milky white semen can be seen being discharged. After 5-10 minutes, the female fish begins to ovulate and chases the male fish, swimming and circling in rapid waves in the breeding institution, which can last for more than 4 hours before turning into low tide. At this time, the water becomes turbid and contains a large number of fertilized eggs.

[0121] After the third feeding, no water is added until the next morning. That is, no water is added before the first feeding time to prevent the sperm and eggs from flowing out with the water.

[0122] In some embodiments, Figure 5 As shown, after the third feeding, no water was introduced, and the seawater was aerated. The aeration amount was based on not generating air waves, so as not to disturb the mating behavior of the lancelets and cause damage to the fertilized eggs.

[0123] S6, after the third feeding in step S5, fertilized eggs or larvae hatched from the fertilized eggs are collected in the seawater, and the larvae hatched from the fertilized eggs are the first generation.

[0124] In some embodiments, seawater is filtered to collect fertilized eggs in the water.

[0125] like Figure 5 As shown, the device also includes a collecting mechanism and a pumping mechanism.

[0126] The collecting mechanism is placed in the breeding mechanism, the top of the collecting mechanism is arranged above the surface of the seawater in the breeding mechanism, the bottom of the collecting mechanism is arranged below the surface of the seawater in the breeding mechanism, and a second filter is arranged at the bottom of the collecting mechanism for collecting fertilized eggs, and the filtered seawater is returned to the breeding mechanism.

[0127] For example, a 150-mesh screen is used as the second filter.

[0128] The flow rate of the water pumped by the pumping mechanism can form a continuous water flow.

[0129] In some embodiments, the flow rate of water pumped by each pumping mechanism is 0.3-0.4 L / min.

[0130] The next morning, take out the collecting mechanism and the pumping mechanism, and then feed for the first time.

[0131] In some embodiments, step S7, starting from the second day in step S5, the feeding time is adjusted three times a day, the interval between each feeding is 6-7 hours, the feeding amount is increased to 1.6-2 times that in step S3, and no water is added after the third feeding until the first feeding on the second day;

[0132] Step S8, after the third feeding in step S6, collecting the fertilized eggs in the seawater, or the larvae obtained by hatching the fertilized eggs.

[0133] In some embodiments, on the day of step S5, although the lancelets show signs of pre-mating behavior, mating behavior does not occur, steps S7 and S8 are still performed until mating behavior occurs.

[0134] Amphioxus only mates at night, so no fertilized eggs are produced during the day and the seawater replacement cycle can continue.

[0135] For example, the specific feeding times are adjusted to 5:30, 11:30, and 17:30.

[0136] Corresponding to the above 3mL, the feeding amount is increased to 5-6mL of algae nutrient solution each time.

[0137] As mentioned above, due to the different development of the gonads of the parents, the mating behavior of different individuals in the group-raised lancelets will vary, forming a relatively continuous period of time. During this period of time, increasing the feeding amount can meet the mating needs of the lancelets. At the same time, feeding in advance can avoid affecting the mating environment of the lancelets, thereby promoting their mating behavior.

[0138] In some embodiments, the method comprises:

[0139] S9, the first generation obtained by hatching the fertilized eggs are cultured to adults and cultured in groups. Each group of first generation offspring corresponds to a group of parent seed preservation. At least two groups of first generation offspring are paired. For each group of first generation offspring, individual lancelets with a body length of 35-55mm, rapid activity, strong sand drilling ability, and full gonads are selected. The paired groups are subjected to steps S2-S6 to obtain the second generation offspring.

[0140] Specifically, each two different groups can be paired to increase the genetic diversity of the next generation.

[0141] For example, n sub-generations can be Group pairing.

[0142] The method for cultivating the first generation includes:

[0143] Seedling cultivation:

[0144] It takes 10-12 hours for the lancelet to go from fertilized eggs to embryos hatching and becoming planktonic larvae.

[0145] After 2-3 days, the larva opens and the first gill slit appears.

[0146] The amphioxus is in the larval stage before metamorphosis is complete.

[0147] Plastic buckets are used as containers for cultivating larvae. The seawater in the plastic buckets is aerated and the cultivation density of the larvae is controlled at 10-40 per milliliter of seawater depending on the size of the larvae.

[0148] In the early and middle stages of the larvae, there is no need to pad sand in the plastic bucket. After the larvae open their mouths, they should be fed with algae nutrient solution in time. Feed twice a day, and replace part of the seawater before feeding. The seawater used for raising seedlings is filtered seawater.

[0149] During the larval stage, the algae nutrient solution fed is mainly golden algae. Before the larvae develop to 6 gill slits, the feeding amount is about 35,000 golden algae / mL seawater each time. During the 6-12 gill slits stage, the feeding amount is about 40,000 golden algae / mL seawater each time.

[0150] In the sub-adult stage, after the metamorphosis and sand drilling, the golden algae can be replaced with flat algae in appropriate amounts. The replacement method is the same as described in the seed preservation method.

[0151] In the early stages of larval cultivation, some larvae may die due to excessive light, so shading is required to avoid direct sunlight.

[0152] Food residues, feces and other impurities deposited in plastic barrels need to be cleaned up in time, otherwise they will seriously affect the survival of the young.

[0153] When cleaning, first pour the clean seawater on the top of the water body together with the larvae floating in the water into another clean plastic bucket. The seawater at the bottom of the plastic bucket can be filtered using a 300-mesh silk sieve to concentrate the dirt, and then transferred to a glass culture dish. Under a dissecting microscope, use a pipette to slowly suck out the dirt, leaving the larvae.

[0154] When the larvae develop to more than 8 gill slits, you can start adding fine sand so that the larvae can dive into the sand in time after metamorphosis. The fine sand used to feed sub-adult lancelets is finer than that of adults and can pass through a 0.15mm metal mesh screen. In order to facilitate cleaning, the sand layer should not be too thick at the beginning and can be controlled below 1cm.

[0155] The mortality rate is very high during the process of nurturing from fertilized eggs to adults. The larvae will die if you are not careful. Appropriate maintenance measures are needed. As the survival and adaptability of the larvae increase after metamorphosis and burrowing into the sand, the mortality rate decreases, the number of the group tends to stabilize, and the survival rate of the offspring until the completion of metamorphosis can be greatly improved.

[0156] The juveniles with too high density should be promptly cultured in separate barrels, and the thickness of the sand layer should be increased to 5-10cm, and the particle size of the sea sand should be increased to 0.5-1mm. At the same time, pay attention to the cleanliness of the sand layer and clean it in time.

[0157] During the subadult stage when the metamorphosis of lancelets is completed and the gonads begin to develop, lancelets live in the sand. When changing the water, put a small amount of sea sand together with the lancelets in batches into a large culture dish, then add an appropriate amount of clean sea water and shake it gently. When the lancelets are exposed above the sand, pour the sea water together with the lancelets into another clean plastic bucket. Repeat this process until all the lancelets are screened out, and then suck away the dirt in the water.

[0158] For subadults over 1 cm in length, the lancelets can be separated from the dirty seawater with the help of a 50-100 mesh sieve.

[0159] The first generation obtained from seed preservation by different breeding institutions is cultivated separately, so as to achieve hybridization during the seed preservation process of the first generation and avoid inbreeding.

[0160] The embodiment of the present disclosure provides a device for preserving the seeds of lancelets, which is used in the above-mentioned method for preserving the seeds, and the device comprises:

[0161] A breeding structure 1, the bottom of which is covered with sea sand 2, and the breeding structure 1 is filled with sea water 3.

[0162] In some embodiments, the breeding structure 1 is provided with a bracket or supporting feet to facilitate the arrangement of the drainage pipeline.

[0163] The bottom of the breeding mechanism 1 is provided with a drain port 4. A plug 41 is installed on the drain port to close the drain port 4.

[0164] A water inlet mechanism 5 is arranged inside the breeding mechanism 1, and the water inlet mechanism 5 is provided with a water inlet 51, and the water inlet 51 is arranged below the water surface of the seawater in the breeding mechanism 1. The water inlet mechanism 5 is arranged along the circumference of the breeding mechanism 1, and the water inlet 51 is oriented in the same direction along the circumference of the breeding mechanism 1.

[0165] The water outlet mechanism 6 is arranged inside the breeding mechanism 1 , and a water outlet 61 is arranged on the upper part of the water outlet mechanism 6 .

[0166] The first filter screen 7 is installed at the bottom of the breeding mechanism 1 , and the first filter screen 7 is arranged at the outer side of the drain port 4 .

[0167] The device further comprises a water storage mechanism 8 , and the water storage mechanism 8 is in communication with the water inlet mechanism 5 .

[0168] The device further comprises an aeration mechanism 9 .

[0169] The aeration mechanism 9 is used to make the seawater in the aquaculture mechanism 1 uniform and increase the dissolved oxygen therein during water outage.

[0170] The aeration mechanism 9 is also used in the cultivation process of larvae.

[0171] The device also includes:

[0172] A collecting mechanism 10, wherein the collecting mechanism 10 is arranged in the aquaculture mechanism 1, the top of the collecting mechanism 9 is arranged above the surface of the seawater in the aquaculture mechanism 1, the bottom of the collecting mechanism 10 is arranged below the surface of the seawater in the aquaculture mechanism 1, and a second filter screen 101 is arranged at the bottom of the collecting mechanism 10;

[0173] The pumping mechanism 11 is used to pump the seawater in the aquaculture mechanism 1 into the collecting mechanism 10 .

[0174] Example 2

[0175] Based on the method described in Example 1, 3 groups of amphioxus parent seed preservation experiments were conducted using PVC plastic barrels. Two parallel experiments were set up for each group of experiments.

[0176] The differences between the three groups of experiments are:

[0177] Group 1: Algae nutrient solution: each feeding amount in step S3, ferric citrate 6×10 -5 ppm / L seawater, urea 1.2×10 -2 ppm / L seawater, potassium dihydrogen phosphate 6×10 -4 ppm / L seawater, 1000 Platymonas / mL seawater.

[0178] Group 2: Algae nutrient solution: each feeding amount in step S3, ferric citrate 6×10 -5 ppm / L seawater, urea 1.2×10 -2 ppm / L seawater, potassium dihydrogen phosphate 6×10 -4 ppm / L seawater, 500 Platymonas / mL seawater, 4500 Chrysophyta / mL seawater.

[0179] Group 3: Algae nutrient solution: the amount of each feeding in step S3, ferric citrate 6×10 -5 ppm / L seawater, urea 1.2×10 -2 ppm / L seawater, potassium dihydrogen phosphate 6×10 -4 ppm / L seawater, 9000 golden algae / mL seawater.

[0180] That is, the algae species are replaced after conversion based on the volume ratio of golden algae to flat algae. 9 golden algae are replaced with 1 flat algae for conversion.

[0181] The remaining seed preservation method steps and seed preservation conditions are the same.

[0182] Observe the amphioxus when they show signs of pre-mating behavior. Since the amphioxus' body is transparent, you can observe the female amphioxus holding eggs.

[0183] The experiment found that in terms of the number of eggs carried by female lancelets, the best effect was achieved by using a nutrient solution containing only golden algae, followed by a nutrient solution containing golden algae and flat algae in a ratio of 1:1. The effect of using a nutrient solution containing only flat algae was not ideal, with female lancelets carrying few eggs, and some even not carrying any eggs.

[0184] Moreover, even though the phenomenon of egg-holding was observed in the third group of experiments, the maturity of the eggs could not be judged from their overall appearance.

[0185] In the second group of experiments, it can be observed that almost all female lancelets can carry eggs, and the overall appearance of the eggs shows that they are of good maturity. In subsequent experiments, seed preservation can be successfully achieved.

[0186] Example 3

[0187] Based on the method described in Example 1, a seed preservation experiment of amphioxus parents was carried out using 6 PVC plastic barrels.

[0188] Algae nutrient solution: each feeding amount in step S3, ferric citrate 6×10 -5 ppm / L seawater, urea 1.2×10 - 2 ppm / L seawater, potassium dihydrogen phosphate 6×10 -4 ppm / L seawater, 500 Platymonas / mL seawater, 4500 Chrysophyta / mL seawater.

[0189] Fertilized eggs were successfully obtained through 6 breeding institutions and hatched to obtain the first-generation larvae, indicating that the seed preservation method provided by the embodiment of the present disclosure can enable the parents to successfully mate and reproduce to obtain the first-generation larvae.

[0190] The first generation obtained from each breeding institution is cultivated in groups, and 10,000-20,000 of them are selected from each group for statistics. After about 11 months, they are cultivated to adults and meet the requirements for seed preservation. At this time, according to counting statistics, the survival rate of the first generation is about 14%. Among them, the survival rate of the first generation from fertilized eggs to sand-boring larvae is about 20%. The survival rate of surviving sand-boring larvae to adults is about 70%.

[0191] Every two groups are paired, and each pair is placed in a breeding institution. The same method is used for seed preservation. The first generation offspring can also mate and reproduce to obtain the second generation offspring. This shows that the first generation offspring obtained by the seed preservation method provided in the embodiment of the present disclosure is relatively strong and can maintain the ability to reproduce and can reproduce to obtain the second generation offspring.

[0192] The second generation of offspring obtained from each breeding institution was cultivated in groups, and six groups were selected. 10,000-20,000 of them were selected from each group for statistics, and they were cultivated to adults after about 11 months and met the requirements for seed preservation. At this time, according to counting statistics, the survival rate of the second generation of offspring was basically the same as that of the first generation, about 14%. Similarly, the survival rate of the second generation of offspring from fertilized eggs to sand-boring larvae was about 20%. The survival rate of surviving sand-boring larvae to adults was about 70%.

[0193] The survival rate of the second generation offspring after being cultivated to adults did not decline, indicating that the seed preservation method provided in the embodiments of the present disclosure can ensure the reproduction quality of lancelets, and the second generation offspring obtained from the artificial secondary reproduction of the parents will not affect the robustness and activity of lancelets.

[0194] In subsequent experiments, the second generation was able to continue to reproduce and produce the third generation, which will not be elaborated here.

[0195] The above description is only a preferred embodiment of the present disclosure and is not intended to limit the present disclosure. For those skilled in the art, the present disclosure may have various modifications and variations. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present disclosure shall be included in the protection scope of the present disclosure.

Claims

1. A method for preserving the seeds of lancelets, characterized in that: The method comprises: S1, collect live lancelets from natural sea areas as parent populations, and select lancelets with fast movement and strong sand drilling ability; S2: The same group of fish were cultured in a seawater environment with sea sand. The thickness of the sea sand was 5-10 cm. The water was continuously circulated and the water was seawater containing algae. The culture density was 6000-12000 fish / m 2 ; S3, feeding algae nutrient solution three times a day, stopping water intake for 2 hours after feeding, and the interval between each feeding was 7-9 hours; S4, drain the cultured seawater twice a day, with an interval of 12 hours between each drainage; S5, if the lancelets were found to have emerged from the sea sand and were swimming in the sea water, or if there was a special smell in the air, they would not be fed with water after the third feeding of the day until the first feeding of the next day; S6, after the third feeding in step S5, collecting fertilized eggs in the seawater, or larvae hatched from the fertilized eggs, the larvae hatched from the fertilized eggs are the first generation; Algae-containing seawater: pH value is 8.3-8.5, ammonia nitrogen mass concentration is less than 0.05 mg / L, dissolved oxygen is higher than 5.5 mg / L, water temperature is 26-32°C, salinity is 19-29, and algae-containing seawater is obtained by filtering seawater from seawater ponds; Algae nutrient solution: each feeding amount in step S3, ferric citrate 6-6.2×10 -5 ppm / L seawater, urea 1.2-1.3×10 -2 ppm / L seawater, potassium dihydrogen phosphate 6-6.2×10 -4 ppm / L seawater, 6000-10000 golden algae / mL seawater, or, replace no more than half of the golden algae with flat algae, and convert based on the volume ratio of golden algae to flat algae.

2. The method according to claim 1, characterized in that Select lancelets with a body length of 35-55mm and full gonads.

3. The method according to claim 1, characterized in that In the algae nutrient solution, golden algae are not replaced by flat algae.

4. The method according to claim 1, characterized in that: Step S4 includes: Before drainage, the sea sand is stirred so that the impurities in the sand float into the sea water and are discharged with the sea water.

5. The method according to claim 1, characterized in that The method further comprises: Step S7, starting from the second day in step S5, the feeding time is adjusted three times a day, the interval between each feeding is 6-7 hours, the feeding amount is increased to 1.6-2 times that in step S3, and no water is added after the third feeding until the first feeding on the second day; Step S8, after the third feeding in step S6, collecting the fertilized eggs in the seawater, or the larvae obtained by hatching the fertilized eggs.

6. The method according to claim 1, characterized in that Step S5 includes: After the third feeding, the seawater was filtered to collect the fertilized eggs in the water.

7. The method according to claim 1, characterized in that Step S5 includes: After the third feeding, the seawater is aerated, and the aeration amount is based on not generating air waves.

8. The method according to claim 1, characterized in that The method further comprises: S9, the first generation obtained by hatching the fertilized eggs are cultured to adults and cultured in groups. Each group of first generation offspring corresponds to a group of parent seed preservation. At least two groups of first generation offspring are paired. For each group of first generation offspring, individual lancelets with a body length of 35-55mm, rapid activity, strong sand drilling ability, and full gonads are selected. The paired groups are subjected to steps S2-S6 to obtain the second generation offspring.

Images