CN111956783B - Artificial spawning induction mixture for largehead black weever and artificial breeding method - Google Patents
Artificial spawning induction mixture for largehead black weever and artificial breeding method Download PDFInfo
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- A61K38/22—Hormones
- A61K38/24—Follicle-stimulating hormone [FSH]; Chorionic gonadotropins, e.g. HCG; Luteinising hormone [LH]; Thyroid-stimulating hormone [TSH]
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; CARE OF BIRDS, FISHES, INSECTS; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K61/00—Culture of aquatic animals
- A01K61/10—Culture of aquatic animals of fish
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; CARE OF BIRDS, FISHES, INSECTS; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K61/00—Culture of aquatic animals
- A01K61/10—Culture of aquatic animals of fish
- A01K61/17—Hatching, e.g. incubators
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/33—Heterocyclic compounds
- A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
- A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
- A61K31/44—Non condensed pyridines; Hydrogenated derivatives thereof
- A61K31/445—Non condensed piperidines, e.g. piperocaine
- A61K31/4523—Non condensed piperidines, e.g. piperocaine containing further heterocyclic ring systems
- A61K31/454—Non condensed piperidines, e.g. piperocaine containing further heterocyclic ring systems containing a five-membered ring with nitrogen as a ring hetero atom, e.g. pimozide, domperidone
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/04—Peptides having up to 20 amino acids in a fully defined sequence; Derivatives thereof
- A61K38/08—Peptides having 5 to 11 amino acids
- A61K38/09—Luteinising hormone-releasing hormone [LHRH], i.e. Gonadotropin-releasing hormone [GnRH]; Related peptides
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P15/00—Drugs for genital or sexual disorders; Contraceptives
- A61P15/08—Drugs for genital or sexual disorders; Contraceptives for gonadal disorders or for enhancing fertility, e.g. inducers of ovulation or of spermatogenesis
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A40/00—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production
- Y02A40/80—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production in fisheries management
- Y02A40/81—Aquaculture, e.g. of fish
Abstract
The invention discloses an artificial spawning induction mixing agent for micropterus salmoides and an artificial breeding method thereof, belonging to the technical field of aquatic animal breeding. The invention provides an artificial induced spawning admixture, which comprises the following components: chorionic gonadotrophin, luteinizing hormone releasing hormone analog No. 2, dilinodone maleate. The artificial spawning induction mixture provided by the invention is used for artificial breeding of the micropterus salmoides, the female fish bred by the artificial spawning induction mixture is high in spawning, the ovum maturity is high, the fertility rate and the hatching rate are both obviously improved, the breeding efficiency of the micropterus salmoides is effectively improved, and the time is saved.
Description
Technical Field
The invention relates to the technical field of aquatic animal cultivation and breeding, in particular to an artificial spawning induction mixing agent for micropterus salmoides parent fish and an artificial breeding method.
Background
The edible weever is beneficial to nourishing liver and kidney, tonifying spleen and stomach, resolving phlegm and relieving cough, and also beneficial to wound healing and promoting brain development of children. In recent years, with the gradual improvement of the quality requirements of consumers on aquatic products, weever has replaced four families of fish, crucian, snakeheads and the like, and becomes an important consumer product on a dining table of people, and the consumer market is expanding continuously. The parent fish and offspring seed supply of the weever can not meet the market demand far, so the price of the weever is always in the rising trend, and the weever has extremely high market value.
The weever culture water area is very wide, and the weever can grow well in common freshwater ponds, freshwater factories, net cages, inland saline-alkali water areas, and has the advantages of excellent environment adaptability, stronger disease resistance, wide feed sources and the like, so that the weever can become an excellent artificial freshwater culture variety in China. The yield of the freshwater aquaculture weever in Guangdong, zhejiang, jiangsu and other places is also considerable. According to the data of Chinese fishery statistics annual book 2019, the fresh water aquaculture yield of the weever in Guangdong province in 2018 reaches 25.84 ten thousand tons, and the position is the first place nationwide. While achieving these favorable results, we noted that female weever is generally mature for 1 year, belonging to a fish with multiple ovulations in one brood. The one-time egg-carrying amount can reach 4-12 ten thousand grains, and the average egg-carrying amount is 6 ten thousand grains. 1-3 ten thousand mature eggs can be discharged in each ovulatory period, and 2-3 times of mature eggs can be discharged in each reproductive period. The egg yield, ovulation yield and ovulation frequency of parent fish are closely related to nutrition and peripheral environment and management. In each spawning period of parent fish, 30% -50% of ova may be blocked or overripe due to insufficient nutrition supply or incapacity of timely ovulation, so that the quality of the ova is reduced, the fertilized ova and the hatching rate of the ova are low, the survival rate of offspring seeds is poor, and the like. Therefore, how to develop an artificial breeding method capable of improving the single ovulation quantity of the micropterus salmoides, increasing the fertility rate and the hatching rate of ova and improving the survival rate of offspring seeds is a technical problem which needs to be solved urgently in the field.
Disclosure of Invention
In order to solve the problems in the prior art, the invention aims to provide the artificial spawning induction mixture, which can obviously improve the ovulation quantity and the ovum quality of the micropterus salmoides and avoid the occurrence of ovum locking.
In order to achieve the above object, the present invention provides the following technical solutions:
the invention provides an artificial spawning induction mixing agent for large-mouth black weever, which comprises the following components: chorionic gonadotrophin 1200-2000 IU/mL, luteinizing hormone releasing hormone analog No. 2 150-220 mug/mL and dyclonone maleate 10-25 mg/mL.
Preferably, the chorionic gonadotrophin in the artificial oxytocin blend is 1500IU/mL, the luteinizing hormone analog No. 2 is 200 μg/mL, and the dyclonone maleate is 10mg/mL.
The invention also aims at providing the application of the oxytocic mixture in artificial breeding of the largehead jewfish.
The invention also aims at providing an artificial breeding method of the micropterus salmoides.
In order to achieve the above object, the present invention provides the following technical solutions:
the invention provides an artificial breeding method of largemouth black weever, which comprises the following steps:
(1) Artificial induced spawning: selecting parent fish, and 15 pm: 00-16: 00 female fish is injected with the oxytocic mixture; placing female fish injected with the mixture and male fish which are not induced to spawn in the same water area for temporary culture, and separating by a net cage; the next morning 6: 00-7: 00 is that the male fish is injected with the oxytocic mixture; after the male fish induced spawning for 3-6 hours, the net cage is removed, so that the female fish and the male fish are fully contacted;
(2) Collecting ovum and sperm of parent fish, artificial insemination, fertilized ovum hatching and larva cultivation;
(3) Culturing the parent fish after ovulation for 8-12 d, then carrying out induced spawning according to the step (1), and carrying out secondary ovulation fertilization.
Preferably, the injection amount of the oxytocic mixture in step (1) is: every 500g of female fish is injected with 0.8-1.2 mL of oxytocic mixture, and the injection quantity of male fish is half of the injection quantity of current female fish.
Preferably, the water temperature for the cultivation of the parent fish is 22-24 ℃ and the water flow speed is 10-20L/min.
Preferably, the parent fish is fed with feed twice daily during the cultivation period, wherein the protein level in the feed is 44-46%, and the grease level is 8-10%.
Preferably, the mixed solution of vitamin E and vitamin A, which is sprayed on each kilogram of feed, is fed for 1 time every 2 days.
Preferably, the above mixed solution is prepared from vitamin E: vitamin A is composed according to the weight ratio of 1:2.
Preferably, the hatching conditions in step (2) comprise: fertilized egg density is 1.5-2.5 ten thousand grains/liter, water flow speed is 2-4L/min, and water temperature is 22-24 ℃.
Preferably, the method comprises the steps of,culturing larvae after the weever is hatched for 6-7 d, wherein the culturing conditions of the larvae in the step (2) are as follows: the density of the seedlings is 8000-10000 tails/m 3 The water flow speed is 5-8L/min, the water temperature is 22-24 ℃, and 60-80 mL/m is added into the water every 3 days 3 Is an EM strain of (A).
Compared with the prior art, the technical scheme of the invention has the following beneficial effects:
(1) According to the invention, by improving parent fish cultivation and selection, the parent fish breeding efficiency is effectively improved, the selected parent fish has high maturity, the female fish has high spawn quantity, and the average spawn quantity is more than 8 ten thousand grains;
(2) The oxytocic and the oxytocic technology provided by the invention can effectively promote gonad development of female fishes and male fishes, improve ovulation quantity and ovulation efficiency of the female fishes, enable the ovulation quantity of the female fishes to be 4-6 ten thousand grains each time, enable the ovulation quantity to be more than 90% of the total egg quantity within 2 ovulation time, and reduce occurrence probability of egg locking and overripening;
(3) The invention provides the oxytocic and the oxytocic technology, which can effectively improve the quality of ova, the fertility rate of the ova is higher than 85%, the hatching rate is higher than 90%, and the survival rate of offspring seeds is higher than 90%.
Detailed Description
The invention provides an artificial spawning induction mixing agent for large-mouth black weever, which comprises the following components: chorionic gonadotrophin 1200-2000 IU/mL, luteinizing hormone releasing hormone analog No. 2 150-220 mug/mL, and dyclonone maleate 10-25 mg/mL. According to the invention, through compounding of the oxytocic substances, the oxytocic mixture is obtained, and after the oxytocic mixture is injected into the parent fish of the micropterus salmoides, the ovulation quantity of the micropterus salmoides can be obviously improved, and the phenomena of ovum locking and overripening are avoided. The ratio is preferably 1500IU/mL of chorionic gonadotrophin, 200 mug/mL of luteinizing hormone releasing hormone analogue No. 2, and 10mg/mL of dyclonone maleate. The chorionic gonadotrophin, luteinizing hormone releasing hormone analogue No. 2 and the diurone maleate can be all commercially available products. In a specific embodiment of the invention, chorionic gonadotrophin, luteinizing hormone releasing hormone analog No. 2, and diurone maleate are purchased from Ningbo second hormone works.
The invention also provides application of the oxytocic mixture in artificial breeding of the largemouth black weever.
The invention also provides an artificial breeding method of the largemouth black weever, which comprises the following steps:
(1) Artificial induced spawning: selecting parent fish, and 15 pm: 00-16: 00 female fish is injected with the oxytocic mixture; placing female fish injected with the mixture and male fish which are not induced to spawn in the same water area for temporary culture, and separating by a net cage; the next morning 6: 00-7: 00 is that the male fish is injected with the oxytocic mixture; after the male fish induced spawning for 3-6 hours, the net cage is removed, so that the female fish and the male fish are fully contacted;
(2) Collecting ovum and sperm of parent fish, artificial insemination, fertilized ovum hatching and larva cultivation;
(3) Culturing the parent fish after ovulation for 8-12 d, then carrying out induced spawning according to the step (1), and carrying out secondary ovulation fertilization.
The invention selects the female parent fish with normal body shape and body color and strong physique and no disease disability to culture the parent fish 1 month before breeding the parent fish. As a preferred implementation mode, the parent fish is put into flowing water with the water temperature of 22-24 ℃ and the water flow speed of 10-20L/min for cultivation, and the effect of stimulating the gonad development of the parent fish is achieved and the reproductive capacity of the parent fish is improved through adjusting the water temperature and the water flow speed. Wherein the water temperature is more preferably 23 ℃, and the water flow speed is more preferably 15L/min. The invention can adopt the conventional cultivation method for other environmental conditions such as illumination, ventilation and the like of parent fish cultivation.
The feed is fed every day during the cultivation period of parent fish, preferably 2 times per day. The protein level of the feed is preferably 44-46%, the grease level is preferably 8-10%, the gonad development level of parent fish is promoted, and the spawning capacity and ovum level are improved. The invention has no limit to the content level of ash and crude fiber in the feed, and the conventional content can be used. The feed of the present invention further preferably has a protein level of 45% and an oil level of 9%. The feed of the invention can be prepared from fish oil: soybean oil: coconut oil is prepared from 50-60 percent: 20-30%: 20-30% of a composition. The feed provided by the invention provides enough nutrition for parent fish through reasonable proportioning of feed nutrition level, and has the effects of promoting gonad development, improving ovum quality and promoting ovulation of parent fish compared with conventional feed special for weever. The fish oil, the soybean oil and the coconut oil are all prepared from products which are commercially available in the field.
In the invention, during the feeding period of the parent fish, 60-100 mg of vitamin E+vitamin A mixed solution is sprayed on each kilogram of feed every 2 days for feeding the parent fish for 1 time, and more preferably 80mg of vitamin E+vitamin A mixed solution is sprayed on each kilogram of feed. The vitamin mixture is prepared from vitamin E: vitamin A is composed according to the weight ratio of 1:2. According to the invention, the quality of the ovum is obviously improved by adding the vitamin mixed solution with a specific proportion, so that the fertilization rate, the hatching rate and the survival rate of the offspring seeds in the later period are improved. In the invention, vitamin E and vitamin A are all commercially available products in the field.
The invention selects the pre-bred female fish which has thick and short body shape, enlarged and soft abdomen, obvious ovarian contour, uniform upper and lower abdomen, elasticity, slightly convex urogenital nipple, ruddy spawning period and 2 holes; the male fish with thin and long body, depressed urine nipple, ruddy urine nipple, 1 hole, slightly pressed abdomen, a small amount of milky semen flowing out and scattered in water is selected for artificial induced spawning. The parent fish selection process is carried out by selecting parent fish with mature gonad according to a conventional method in the field.
In the artificial spawning induction stage, the spawning induction mixture is respectively injected into parent fish bodies, the injection part is the inner side of the ventral fin of the fish body, the needle is obliquely put down, and the needle depth is 1-1.2 cm, preferably 1.1cm; the injection amount of the oxytocic mixture is as follows: each 500g of female fish is injected with 0.8-1.2 mL, more preferably 0.9-1 mL, and the injection quantity of the male fish is half of that of the female fish.
The invention is 15 pm: 00-16: and (3) carrying out injection of the oxytocic mixture on the female fish at the time of 00. Since weever usually spawns in the evening with weaker light, the single injection effect time of the oxytocic mixture is 24-25 hours, therefore, the invention finds that the weever is selected at 15 pm: 00-16: the injection at the time of 00 is more favorable for the smooth ovulation of female fishes compared with other time, and can further improve the single ovulation quantity without influencing the quality of ova. The injection time of the male fish is 15-17 hours after the injection of the female fish, and the injection time of the male fish is 6 a.m. in the following day: 00-7: 00, the synchronous development and maturation of the sperm of the male fish and the ovum of the female fish can be ensured, and the fertilization rate can be effectively improved in the artificial insemination process.
According to the invention, female fish injected with oxytocin and male fish which are not induced to spawn are placed in the same water area for temporary culture, and are separated by a net cage, and the female fish is temporarily cultured in the net cage. The net cage is placed in the water body, so that the male fish and the female fish can live in the same water body, the male fish and the female fish are close to each other and are not affected by each other, and the male fish can release some sexual signals in the water body, so that sexual maturity of the female fish is facilitated. 3-6 hours after the oxytocin is injected into the male fish, the net cage is removed, so that the female fish and the male fish are fully contacted, and the net cage removing time is more preferably 4 hours after the oxytocin is injected into the male fish. The invention discovers that the arrangement of the net cage can obviously improve the single ovulation quantity of female fish compared with a method for solely temporarily breeding parent fish. The invention has no special requirement on the net cage material, and the mesh size only needs to ensure that parent fish does not pass through.
According to the invention, after the female fish is injected with the oxytocic mixture for 24-26 hours, the abdomen of the female fish is gently extruded, and the ovum is collected; after the male fish is injected with the oxytocic mixture for 9-11 hours, adopting a sucking mode, lightly wrapping gonadal tissues by using a rubber head dropper, and lightly sucking out semen; adding 2-3mL of 0.7% physiological saline into 10000 eggs to promote fertilization holes to open, adding 0.2-0.4 mL of semen, adding 3-5 mL of water to activate fertilized eggs, closing fertilization holes, stirring for 20-30 s, and completing fertilization process to obtain fertilized eggs.
The invention discovers that in the fertilized egg hatching process, the fertilized egg density is regulated to be 1.5-2.5 ten thousand grains/liter, the water flow speed is controlled to be 2-4L/min, the water temperature is controlled to be 22-24 ℃, a good water body environment can be provided for the fertilized egg, the hatching rate of the fertilized egg is effectively improved, and the weever is ensured to start hatching out a membrane after 70-72 hours. The water flow speed is more preferably 3L/min, and the water temperature is 23-24 ℃. Other environmental conditions such as illumination, ventilation and the like in the fertilized egg hatching process can be conventional cultivation methods, and the fertilized egg hatching method is not limited.
After the weever is hatched for 6-7 days,fishing out the seedlings, and putting the seedlings into a seedling cultivation pool, wherein the density of the seedlings is 8000-10000 tails/m 3 Preferably 9000 tail/m 3 Using artemia as an initial bait; continuously inflating during the cultivation period, controlling the water temperature at 22-24 ℃ and the water flow speed at 5-8L/min, changing water for 1/3 every 3d, and ensuring the fresh water environment by the natural light period, wherein the temperature difference between the water before and after changing water is not more than 0.5 ℃. The invention has no special requirements on the selection of the initial feed for the offspring seeds and the offspring seed cultivating pool, and can also be replaced by the conventional initial feed, cultivating barrels and the like.
60-80 mL/m of the seed culture pond is added every 3 days 3 Preferably 70mL/m 3 Is an EM strain of (A). The added amount of EM bacteria can better improve the beneficial bacteria composition of the water body, promote the intestinal development of the micropterus salmoides, and further improve the survival rate of the offspring seeds.
According to the method, the female fish after ovulation is continuously cultivated for 8-12 days, preferably 10 days, according to the parent fish cultivation method, then the spawning induction process is carried out, the female fish egg nest is basically emptied after two times of ovulation, the two times of ovulation accounts for more than 90% of the total spawning quantity of the female fish, the egg maturity is high, the fertility rate and the hatching rate are obviously improved, the breeding efficiency of the female fish is effectively improved, and the time is saved.
The technical solutions of the present invention will be clearly and completely described in the following in connection with the embodiments of the present invention. It will be apparent that the described embodiments are only some, but not all, embodiments of the invention. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
In the specific embodiment of the invention, the largehead jewfish parent fish is 'you weever No. 2', which is purchased from Zhujiang aquatic institute of China aquatic science institute, but is not limited to the technical scheme of the invention.
Example 1
An improved technique for cultivating and artificially breeding the parent fish of largemouth black bass comprises the following steps:
step 1, breeding parent fish for 1 month, and selecting female parent fish with normal body shape and color and strong physique and without disease disability. And (3) respectively placing the selected parent fish into flowing water with the water temperature of 24 ℃ and the water flow speed of 15L/min. During the cultivation period, the feed with the protein level of 44% and the feed grease level of 10% (the ratio of fish oil: soybean oil: coconut oil is 60%:20%: 20%) was fed 2 times per day. Every 2 days, 100mg of vitamin E+vitamin A mixed solution (1:2) is sprayed on each kilogram of feed to feed parent fish for 1 time, so that gonadal development and maturation are accelerated. Meanwhile, the parent fish of the largehead jewfish cultured conventionally is selected as a control group to feed special feed for the largehead jewfish produced by Zhejiang Xintianen aquatic feed limited company, wherein the crude protein content is 45%, the fat is 10%, the crude fiber is 5% and the crude ash content is 16%.
Step 2, selecting 10 tails of the pre-bred female fish, wherein the shapes of the tails are thicker and shorter, the abdomen is enlarged and soft, the outline of the ovary is obvious, the upper abdomen and the lower abdomen are uniform in size and elastic, the urogenital nipple is slightly convex, the spawning period is ruddy, and the tails are provided with 2 holes. Female fish body: the body width is 2:1. the male fish 4 tail is thin and long in shape, the urogenital nipple is concave and ruddy, only one hole is formed, and a small amount of milky semen flows out from the lightly pressed abdomen and is dispersed in water.
Step 3, the experimental group adopts an artificial oxytocic mode to respectively prepare a mixture of the oxytocic drug chorionic gonadotrophin 1500 IU/mL+luteinizing hormone releasing hormone analogue No. 2 200 mug/mL+dyclonone maleate 10mg/mL. Each 500g female fish is injected with 0.8ml of the above-mentioned oxytocic mixture. The injection time was 15 pm: 00, the injection part is the inner side of the ventral fin of the fish body, the needle is obliquely lowered, and the needle depth is 1cm. The following morning, the male fish 6:00 injection of the oxytocic mixture, the injection mode is the same as that of female fish, and the injection metering is halved. The control group adopts natural breeding, and the male-female ratio is 5:2, 10 female fish and 4 male fish, and placing the male and female fish at 8m 3 In the temporary raising barrel, a palm net sheet is placed on the bottom edge of the barrel.
Step 4, placing the female fish injected with oxytocin and the male fish without oxytocin into the same circular temporary rearing barrel with the volume of 8m 3 . Female fish temporarily cultured in barrel with specification of 2m 3 Is isolated from the male fish. After the oxytocin is injected into the male fish for 4 hours, the net cage is removed, so that the female fish and the male fish are fully contacted, and ovulation is promoted.
Step 5, after 24 hours of injection, the effect time of the male fish is reached after 9 hours. At this time, it is obviously observed that the male fish surrounds the female fish, the mouth is continuously used for contacting the abdomen and the reproduction hole of the female fish, and the tail part is continuously used for beating the female fish, so that the male fish expects a breeding signal matched with the female fish. Lightly squeezing the abdomen of female fish, and collecting ovum with stainless steel basin; the male fish adopts a sucking mode, and after neutral gland tissues are lightly wrapped by a rubber head dropper, semen is lightly sucked out. 0.2mL semen is added into each 10000 eggs, 5mL water is added for activating, and stirring is carried out for 30s, thus completing the fertilization process. After the fertilized eggs are de-bonded by yellow mud, the fertilized eggs are placed in hatching barrels, 20 ten thousand eggs are placed in each hatching barrel, the water flow speed is 3L/min, and the water temperature is 23 ℃. The fertilized eggs of the control group are hatched with conventional palm net sheet running water, fish fries are collected after hatching for 7 days, and the number of the fish fries is counted.
Step 6, culturing the female fish after ovulation for 10 days according to the operation of step 1, and then carrying out the induced spawning process. The control group was further cultured for 10 days, and then further cultured.
Table 1 weight of parent fish in experimental and control groups
As can be seen from table 1, after the parent fish cultivation, the average body weight of the experimental group 10 females was 846.33g, the average body weight of the 4 males was 682.48g, the average body weight of the control group 10 females was 841.29g, and the average body weight of the 4 males was 713.52g, indicating that there was no significant difference in body weight (P > 0.05) between the experimental group and the control group females and males.
TABLE 2 comparison of spawning amount, fertility Rate and hatchability of female fish in Experimental group and control group
9 females in the 10 experimental groups successfully induced spawning in the first ovulation period, and the first and second ovulation amounts, fertilization rate and hatching rate of the 9 females are counted respectively. In the control group, 10 female fish were laid in succession within 3 days. As can be seen from table 2, the experimental group had higher spawning numbers twice, fertilization rates, and hatchability than the control group.
Example 2
An improved technique for cultivating and artificially breeding the parent fish of largemouth black bass comprises the following steps:
step 1, breeding parent fish for 1 month, and selecting female parent fish with normal body shape and color and strong physique and without disease disability. And (3) respectively placing the selected parent fish into flowing water with the water temperature of 22 ℃ and the water flow speed of 10L/min. During the cultivation period, feeds with protein level of 45% and feed grease level of 9% (fish oil: soybean oil: coconut oil in a ratio of 50%:20%: 30%) were fed 2 times per day. Every 2 days, 100mg of vitamin E+vitamin A mixed solution (1:2) is sprayed on each kilogram of feed to feed parent fish for 1 time, so that gonadal development and maturation are accelerated. Meanwhile, conventionally cultured micropterus salmoides parent fish is selected as a control group. Meanwhile, the parent fish of the largehead jewfish cultured conventionally is selected as a control group to feed special feed for the largehead jewfish produced by Zhejiang Xintianen aquatic feed limited company, wherein the crude protein content is 45%, the fat is 10%, the crude fiber is 5% and the crude ash content is 16%.
Step 2, selecting 8 tails of pre-bred female fish, wherein the shapes of the tails are thicker and shorter, the abdomen is enlarged and soft, the outline of the ovary is obvious, the upper abdomen and the lower abdomen are uniform in size and elastic, the urogenital nipple is slightly convex, the spawning period is ruddy, 2 holes are formed in the urogenital nipple, and the female fish body is high: the body width is preferably (3:1). The male fish 4 tail is thin and long in shape, the urogenital nipple is concave and ruddy, only one hole is formed, and a small amount of milky semen flows out from the lightly pressed abdomen and is dispersed in water.
Step 3, preparing a mixture of the oxytocic drug chorionic gonadotrophin 1500IU/mL, the luteinizing hormone releasing hormone analogue No. 2 200 mug/mL and the dyclonone maleate 10mg/mL respectively. Each 500g female fish is injected with 0.8mL of the above-mentioned oxytocic mixture. The injection time was 16 pm: 00, the injection part is the inner side of the ventral fin of the fish body, the needle is obliquely arranged, and the needle depth is 1.1cm. Male fish 7 in the morning of the next day: 00 injection of the oxytocic mixture, the injection mode is the same as that of female fish, and the injection metering is halved. And naturally breeding 8 tails of female fish and 4 tails of male fish in a control group.
Step 4, placing the female fish injected with oxytocin and the male fish without oxytocin into the same round temporary rearing barrel with the volume of 4m 3 . Female fish temporarily cultured in barrel with specification of 1m 3 Is isolated from the male fish. After the oxytocin is injected into the male fish for 4 hours, the net cage is removed, so that the female fish and the male fish are fully contacted, and ovulation is promoted. Female fish and male fish of the control group are placed in a 4m3 cultivation barrel for cultivation, and brown meshes are placed on the bottom edge of the barrel.
Step 5, after 26 hours of injection, the effect time of the male fish is reached after 11 hours. At this time, it is obviously observed that the male fish surrounds the female fish, the mouth is continuously used for contacting the abdomen and the reproduction hole of the female fish, and the tail part is continuously used for beating the female fish, so that the male fish expects a breeding signal matched with the female fish. Lightly squeezing the abdomen of female fish, and collecting ovum with stainless steel basin; the male fish adopts a sucking mode, and after neutral gland tissues are lightly wrapped by a rubber head dropper, semen is lightly sucked out. 0.2mL of semen is added into each 10000 eggs, 5mL of water is added for activating, and stirring is carried out for about 30 seconds, thus finishing the fertilization process. After the fertilized eggs are de-bonded by yellow mud, the fertilized eggs are placed in hatching barrels, 20 ten thousand eggs are placed in each hatching barrel, the water flow speed is 3 liters/min, and the water temperature is 24 ℃. The fertilized eggs of the control group are hatched with conventional palm net sheet running water, fish fries are collected after hatching for 7 days, and the number of the fish fries is counted.
Step 6, after hatching for 72 hours, the weever starts to grow the fry and starts to grow the membrane, and after 7 days, the weever is in a parallel swimming state, and the fry is collected.
TABLE 3 weight of parent fish in experimental and control groups
As can be seen from table 3, after the parent fish cultivation, the average body weight of the experimental group 8 females was 756.33g, the average body weight of the 4 males was 693.26g, the average body weight of the control group 8 females was 765.23g, and the average body weight of the 4 males was 685.18g, indicating that there was no significant difference in body weight (P > 0.05) between the experimental group and the control group females and males.
TABLE 4 comparison of the total amount of ovulation and the egg quality of the female fish in the experimental and control groups
As can be seen from Table 4, the number of ovulations of each female fish in the experimental group is 8.7-11.3 ten thousand, and the number of ovulations of each female fish in the control group is 5.3-7.2 ten thousand, which indicates that the total number of spawning of each female fish in the experimental group is significantly higher than that in the control group. Meanwhile, the locking rate of the oocyte in the ovum of the experimental group is 9.6-13.8%, and the locking rate of the oocyte in the control group is 15.3-19.2%, which is obviously higher than that of the experimental group. The twice fertilization rate and the twice hatching rate in the experimental group are respectively 86.3-92.5% and 93.4-94.8%, and the twice fertilization rate and the twice hatching rate in the control group are respectively 67.2-72.8% and 74.1-75.5%, which are both remarkably lower than those of the experimental group.
TABLE 5 Total emergence and survival rates of seedlings in the experimental and control groups
As shown in Table 5, the emergence rate of the seedlings bred according to the improved technology can reach 160 ten thousand, which is far higher than 76 ten thousand in the control group, the survival rate of the seedlings can reach 90.3-91.2%, which is obviously higher than 82.5-84.5% of the control group. The improvement method is shown to greatly improve the parent fish utilization and breeding efficiency of the micropterus salmoides, promote the high-quality offspring seed supply of the micropterus salmoides and accelerate the industrialized development.
Example 3
In order to explore the efficacy of the oxytocic mixture obtained by compounding the oxytocic drug, the following single-factor experiment is carried out:
according to the parent fish cultivation method in the experimental group 1, parent fish cultivation and selection are carried out, 32 female fish and 16 male fish with the same specification and the same cultivation time are selected, the parent fish cultivation method is randomly divided into four groups, namely an experimental group 1, an experimental group 2, an experimental group 3 and a control group, 8 female fish and 4 male fish in each group are selected, and artificial induced spawning is carried out on the experimental groups 1-3: the oxytocic drug selected in the experimental group 1 is a mixture of 1500IU/mL of chorionic gonadotrophin and 200 mug/mL of luteinizing hormone releasing hormone analogue No. 2 and 10mg/mL of diolone maleate; the oxytocic drug selected in the experimental group 2 is chorionic gonadotrophin 1500IU/mL; the oxytocin drug selected in experimental group 3 was luteinizing hormone-releasing hormone analogue No. 2 200 μg/mL. The injection modes of the oxytocic drug or the oxytocic mixture are as follows: every 500g female fish is injected with 0.8mL of the oxytocic mixture, and the injection time is 16 pm: 00, the injection part is the inner side of the ventral fin of the fish body, the needle is obliquely arranged, and the needle depth is 1.1cm. Placing female fish injected with the mixture and male fish which are not induced to spawn in the same water area for temporary culture, and separating by a net cage; male fish 7 in the morning of the next day: 00 injection of the oxytocic mixture, the injection mode is the same as that of female fish, and the injection metering is halved. The experiment of 3 groups only comprises that the injected oxytocic drugs are different, and other culture conditions are the same. The parent fish in the control group did not inject the oxytocic or oxytocic cocktail, but maintained the same regimen as the experimental group. There was no significant difference in body weight among the groups (see Table 6 for details).
TABLE 6 weight of parent fish in experimental and control groups
The first ovulation amount and the second ovulation amount (see Table 7 for details) of four groups of females were counted, respectively, to show that: the first ovulation amount of the experimental group 1 can reach 3.8-4.2 ten thousand grains, the second ovulation amount can reach 4.2-4.8 ten thousand grains, and the total amount of single ovulation and double ovulation in the experimental group 1 is obviously higher than that of other experimental groups and control groups.
TABLE 7 amount of ovulation twice for experimental and control groups
Example 4
To explore the effect of the feed on the ovulation time and oocyte development of micropterus salmoides during the cultivation of parent fish in the present invention, the following single-factor experiments were performed:
the method comprises the steps of selecting 32 female fish and 16 male fish with the same specification and similar development, randomly dividing the female fish and the male fish into four groups, namely an experimental group 1, an experimental group 2, an experimental group 3 and an experimental group 4, respectively, wherein 8 female fish and 4 male fish in each group are fed with different feeds during parent fish cultivation: feeding the feed with the protein level of 45% and the feed grease level of 9% (fish oil: soybean oil: coconut oil in the proportion of 50%:20%: 30%) for 2 times per day in experimental group 1, and feeding parent fish 1 time by spraying 80mg of vitamin E+vitamin A mixed solution (1:2) on each kilogram of feed every 2 days; the feed in the experimental group 2 is the same as the feed in the experimental group 1, the feeding times are 2 times per day, but the feed is fed to parent fish 1 time by spraying 80mg of vitamin E on each kilogram of feed every 2 days; the experimental group 3 is fed with the special feed (with 45% of crude protein, 10% of fat, 5% of crude fiber and 16% of crude ash) for the micropterus salmoides produced by Zhejiang Xin Tianen aquatic feed limited company for 2 times every day; in the experimental group 4, the feed is the same as that in the experimental group 3, the feeding times are also 2 times per day, but the parent fish is fed 1 time by spraying 80mg of vitamin E+vitamin A mixed solution (1:2) on each kilogram of feed every 2 days. The experimental groups 1 to 4 are different in feed only fed during parent fish cultivation, the rest cultivation conditions are the same, and the modes of post artificial spawning promotion and the like are the same, and are the artificial breeding modes of the experimental group in the embodiment 1. Statistical analysis of ovulation time and oocyte development status of micropterus salmoides in four groups of experiments (see table 8) shows that: the first ovulation time of the largemouth bass in the experimental group 1 is 24-28 days, which is shorter than that of the experimental group 2-3, the interval time between the two ovulations in the experimental group 1 is 6-8 days, which is obviously shorter than that of the experimental group 2-4, and the locking rate of the oocyte in the experimental group 1 is only 9.3-15.3% which is lower than that of the experimental group 2-4. Namely, after the feed formula and the feed additive in the experimental group 1 are combined, the premature ovulation of the micropterus salmoides can be obviously promoted, the period of the two ovulation intervals is shortened, and the oocyte locking rate can be obviously reduced.
TABLE 8 influence of different feed additive groups on ovulation time and oocyte development of micropterus salmoides
The foregoing is merely a preferred embodiment of the present invention and it should be noted that modifications and adaptations to those skilled in the art may be made without departing from the principles of the present invention, which are intended to be comprehended within the scope of the present invention.
Claims (2)
1. The artificial breeding method of the largemouth black weever is characterized by comprising the following steps:
(1) Artificial induced spawning: selecting parent fish, and 15 pm: 00-16: 00 female fish injection of an oxytocic mixture; placing female fish injected with the mixture and male fish which are not induced to spawn in the same water area for temporary culture, and separating by a net cage; the next morning 6: 00-7: 00 is a male fish injection induced spawning mixture; after the male fish induced spawning for 3-6 hours, the net cage is removed, so that the female fish and the male fish are fully contacted;
(2) Collecting ovum and sperm of parent fish, artificial insemination, fertilized ovum hatching and larva cultivation;
(3) Culturing the ovulated parent fish for 8-12 d, then carrying out induced spawning according to the step (1), and carrying out secondary ovulation fertilization;
the oxytocic mixture consists of chorionic gonadotrophin 1200-2000 IU/mL, luteinizing hormone releasing hormone analogue No. 2 150-220 mug/mL and dyclonone maleate 10-25 mg/mL;
the injection quantity of the oxytocic mixture is as follows: injecting 0.8-1.2 mL of the oxytocic mixture into each 500g of female fish, wherein the injection quantity of the male fish is half of the injection quantity of the current female fish;
the water temperature for the cultivation of the parent fish before breeding is 22-24 ℃ and the water flow speed is 10-20L/min;
feeding feed twice daily during the cultivation period of parent fish, wherein the protein level in the feed is 44-46% and the grease level is 8-10%;
spraying 60-100 mg of mixed solution of vitamin E and vitamin A on each kilogram of feed every 2 days, and feeding for 1 time; the mixed solution is prepared from vitamin E: vitamin A is composed according to the weight ratio of 1:2;
the hatching conditions include: fertilized egg density is 1.5-2.5 ten thousand grains/liter, water flow speed is 2-4L/min, water temperature is 22-24 ℃, and larva cultivation is carried out after weever hatching out a membrane for 6-7 d, wherein the cultivation conditions are as follows: the density of the seedlings is 8000-10000 tails/m 3 The water flow speed is 5-8L/min, the water temperature is 22-24 ℃, and 60-80 mL/m is added into the water every 3 days 3 Is an EM strain of (A).
2. The method for artificial propagation of micropterus salmoides according to claim 1, wherein said oxytocin cocktail consists of 1500IU/mL chorionic gonadotrophin, 200 μg/mL luteinizing hormone analog No. 2 and 10mg/mL dion maleate.
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