CN112472354A - Artificial insemination method for fertilized fish sebastes schlegeli hilgendorf in ying and building - Google Patents
Artificial insemination method for fertilized fish sebastes schlegeli hilgendorf in ying and building Download PDFInfo
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- 238000000034 method Methods 0.000 title claims abstract description 61
- 241001582957 Sebastes schlegelii Species 0.000 title claims abstract description 60
- 241000251468 Actinopterygii Species 0.000 claims abstract description 140
- 238000009395 breeding Methods 0.000 claims abstract description 54
- 230000001488 breeding effect Effects 0.000 claims abstract description 54
- 239000003085 diluting agent Substances 0.000 claims abstract description 27
- 230000019100 sperm motility Effects 0.000 claims abstract description 11
- 230000001276 controlling effect Effects 0.000 claims abstract description 5
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61D—VETERINARY INSTRUMENTS, IMPLEMENTS, TOOLS, OR METHODS
- A61D19/00—Instruments or methods for reproduction or fertilisation
- A61D19/02—Instruments or methods for reproduction or fertilisation for artificial insemination
- A61D19/027—Devices for injecting semen into animals, e.g. syringes, guns, probes
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; 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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- 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
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Abstract
The invention provides an artificial insemination method for fertilized fish sebastes schlegeli hilgendorf in ying and body; the method comprises the steps of preparing a sebastes schlegeli sperm diluent, measuring sperm motility, artificially inseminating female parent fish, regulating and controlling breeding environment after insemination and counting insemination success rate. The method has the significance that the insemination process of female parent fish can be completed quickly by obtaining a small amount of sperms in the mating season of sebastes schlegeli hilgendorf female and male parent fish, the insemination success rate of the female parent fish is over 91 percent, the problem that the mating rate is low in the mating process due to improper condition control of indoor artificial breeding of the parent fish is solved, the defect that wild or net cage pregnant female fish is adopted for breeding at present is overcome, the full artificial breeding and fry batch production of sebastes schlegeli can be realized, and technical support is provided for the breeding, breeding and proliferation releasing of sebastes schlegeli.
Description
Technical Field
The invention belongs to the technical field of aquaculture marine fish breeding; in particular to an artificial insemination method of sebastes schlegeli hilgendorf fertilized fish in ying.
Background
Sebastes schlegeli (sebastes schlegeli;)Sebastes schlegelii) Also known as Hei , the fish is of the genus Sebastes of Semicifu sebastoidea, Sebastes of Semicidae, Sebastes sebastoidea, is mainly distributed in yellow, Bohai sea and eastern sea areas in China, is also commonly found in the Korean peninsula, Japan, Huocheck sea and the like, and is offshore cold-temperature bottom rocky fish. The ecological breeding cage has the breeding characteristics of strong disease resistance, coarse food intake, natural overwintering and the like, and has the nutritional values of tenderness, delicious taste, rich nutrition and the like, and the two advantages promote the ecological breeding cage to become an important cage breeding object emerging in coastal areas in northern China. Sebastes schlegeli is an oviparous reproduction habit of fertilization in ying and embryo development in vivo, the developmental maturity of female and male glands is asynchronous, male sperms meet with female fishes after sexual maturity in about 11 months every year, at the moment, female ovaries are not sexually mature, sperms are stored in the female fishes, fertilized in about 4 months in spring next year after low temperature period in winter and along with the developmental maturity of the ovaries of the female fishes, and the larval fishes are produced after the in vivo development of embryos in about 1 month.
Sebastes schlegeli hilgendorf has asynchronous gonad developmental maturation, special mating, sperm storage mode, fertilization mode and in-vivo development different from those of oviparous fishes, and cannot simultaneously obtain sexually mature sperms and ova and complete artificial insemination mode through in-vitro fertilization. The development of the full artificial breeding is seriously hindered. At present, two artificial breeding modes of sebastes schlegeli hilgendorf mainly exist, firstly, wild fish in a sea area or cage culture brood parent fish in a breeding season are captured for indoor artificial breeding, and the problems of unstable sources of the brood parent fish, high transportation difficulty, high stress, easy abortion, high yield and mortality rate of the brood fish and the like exist; also has the disadvantages of destroying wild natural resources and biodiversity, etc. Secondly, a mode of indoor mating reproduction of the parent fish is adopted, and if the mating environment is improperly controlled, the problems of low mating rate, low fertilization farrowing rate and the like exist. At present, an effective method for artificial insemination of sebastes schlegeli hilgendorf still does not exist at home and abroad, and quantitative large-scale production of artificial breeding offspring seeds cannot be realized, so that the artificial breeding and artificial propagation of the sebastes schlegeli hilgendorf are realized. The seedling cultivation and the net cage culture industry bring difficulties. Therefore, the method provides a new method for the next step of artificial breeding, selective breeding and cultivation by establishing the artificial insemination method for in vivo fertilization of sebastes schlegeli hilgendorf, and also provides a reference for the artificial breeding of other fish spawning seawater. Solves the problems of low natural mating rate and fertility rate or industrial mating rate and farrowing rate caused by the change of conditions such as sea area temperature and the like.
Disclosure of Invention
The invention aims to provide the artificial insemination method of the fertilized fish sebastes schlegeli hilgendorf in a nutritive cube, the insemination of female parent fish can be completed by acquiring sperms of male fish in the mating season, the operation is simple and rapid, the process of natural mating is simulated and replaced, and the fertilization and spawning rate of the female parent fish is obviously improved.
The invention is realized by the following technical scheme:
the invention relates to an artificial insemination method of sebastes schlegeli hilgendorf fish fertilized in ying, which comprises the following steps:
step 1, preparing a sperm diluent and measuring sperm motility;
step 2, artificial insemination of female parent fish;
step 3, regulating and controlling the breeding environment after insemination;
and 4, counting the success rate of spring insemination.
Preferably, in step 1, the preparation of sebastes schlegeli sperm diluent and the determination of sperm motility specifically comprise:
dissecting the male parent fish to obtain a spermary, weighing and placing the spermary into a culture dish, and weighing the spermary: adding the diluent into the mixture according to the volume of 1:1-1: 1.5; shearing spermary with scissors or placing in a container to break the spermary, filtering tissue fragments with a filter screen for 2 times, sucking sperm diluent and placing in a centrifuge tube for use;
and (3) observing the sperm motility of the sperm diluent under an optical microscope, wherein the sperm diluent with the motility of more than 95 percent is used for artificial insemination materials for standby.
Preferably, in step 2, the artificial insemination of the female parent fish is specifically: anaesthetizing female parent fish with a live fish anesthetic MS-222 (manufacturer: Sigma Aldrich company, USA) with the concentration of 120-.
Preferably, in step 3, the post-insemination culture environment regulation is specifically: the water temperature is controlled to be above 9 ℃, so that great fluctuation is avoided, and flowing water is oxygenated for cultivation; starting night illumination cultivation and nutrition enhancement one month before farrowing, and stopping feeding one week before farrowing; necessary breeding work such as feeding and excrement and residual bait removing is eliminated, illumination and artificial workshop movement are reduced, and a quiet environment is created for the breeding work.
Preferably, in step 4, the statistics of insemination success rate specifically include: in order to avoid damaging the female parent fish, the insemination success rate of the female parent fish is measured according to the situation that whether the female parent fish farrows in spring, the parent fish farrows 4-5 months next year, the tail number of the female parent fish which can farrow is recorded, and the insemination success rate is calculated.
Preferably, the male parent fish is a male parent fish which is grown in a factory domestication sex maturity mode above 3 years old, and the spermary is mature in 11 months per year.
Preferably, the diluent is urine of male parent fish, and the urine is obtained by inserting a 2 mL syringe needle into the bladder to suck the urine when the male parent fish is dissected, and storing the urine in a centrifuge tube for later use.
Preferably, the sebastes schlegeli sebastes female parent fish is a sexually mature female of age 4 or more.
Preferably, the sperm diluent is sucked in a process of avoiding generating air bubbles, and the genital pore is pressed by fingers for 3-5 s immediately after the injection is finished, so that the semen is prevented from flowing out.
Preferably, the formula for calculating the insemination success rate is the tail number of the farrowing female parent fish/the tail number of the artificial insemination female fish multiplied by 100%.
The invention has the following advantages:
(1) the invention relates to an artificial insemination method of sebastes schlegeli hilgendorf fertilized fish in ying and ying; an artificial insemination method of important oviparous economic fish sebastes schlegeli hilgendorf is established, the method replaces the direct collection of wild or cage-cultured brood parent fish for indoor artificial breeding, and solves the problems that the source of the brood parent fish is unstable, the transportation difficulty is high, the cost is increased, the transportation stress is easy to cause the abortion and death of the parent fish, the survival rate of the larval fish is low, the deformity rate is high and the like; the method has the advantages that the female parent fish with high fertility rate and high spawning rate is obtained, the full-manual fertilization of sebastes schlegeli hilgendorf is achieved, the significance of the method is that the fertilization process of the female parent fish can be completed quickly by obtaining a small amount of sperms in the mating season of the female and male parent fish of sebastes schlegeli, and the success rate of female insemination is over 91 percent. The method solves the problem that the mating rate is low in the natural mating process due to improper condition control of indoor artificial breeding parent fish, thoroughly overcomes the defect of breeding by adopting wild or net cage pregnant fish, can realize the full artificial breeding and fry batch production of sebastes schlegeli hilgendorf, and also provides technical support for the breeding, breeding and proliferation releasing of sebastes schlegeli hildorsali.
The whole chain is bred, the blank of the field is filled, and technical conditions are provided for large-scale breeding and breeding of seedlings.
(2) The invention relates to an artificial insemination method of sebastes schlegeli hilgendorf fertilized fish in ying and ying; by establishing an artificial insemination method of sebastes schlegeli hilgendorf, the insemination success rate of sexually mature female parent fish is more than 91 percent; the method has the advantages of feasible production practice, simple and easy operation, low cost and high efficiency, and can finish the fertilization work of even hundreds of female parent fishes by only using a few male fishes with sexual maturity.
(3) Compared with the indoor natural mating method, the artificial insemination method for the fertilized fish sebastes schlegeli hilgendorf in the ying body, disclosed by the invention, avoids the interference of culture conditions such as artificially controlled water temperature, illumination, water flow, sound and the like in the mating period, greatly improves the insemination success rate, further improves the fertilization rate and ensures the smooth breeding of the offspring seeds in the next year.
(4) According to the artificial insemination method for sebastes schlegeli hilgendorf fertilized fish in a nutrient body, the phenomenon of ' one male and more female ' and the phenomenon of one male and more female ' exist in the free mating of sebastes schlegeli hilgendorf, so that the trouble of family identification exists when a breeding family is constructed, and the method provides convenience for the construction of the family of sebastes schlegeli hildorf and the breeding of new lines and varieties.
Detailed Description
The present invention will be described in detail with reference to specific examples. It should be noted that the following examples are only illustrative of the present invention, but the scope of the present invention is not limited to the following examples.
Example 1
The embodiment relates to an artificial insemination method of sebastes schlegeli hilgendorf fertilized fish in ying, which comprises the following specific steps:
step 1, obtaining sperm diluent of male parent fish of sebastes schlegeli hilgendorf:
11 months 8-10 days in 2017, selecting industrially cultured male parent fish of sebasticises schlegeli of more than 4 and 3 ages in Nicotai Huahaiyang science and technology Limited, dissecting the male fish, inserting the male fish into a bladder by using a 2 mL syringe needle to absorb urine, and storing the urine in a centrifugal tube as a sperm diluent for later use; acquiring a spermary, weighing the spermary, putting the spermary into a culture dish, and then weighing the spermary according to the weight of the spermary: the diluent is added rapidly according to the volume ratio of 1:1-1:1.5, the spermary is cut up by scissors or is placed in a container to be ground, a 300-mesh filter screen (a silk screen) is used for filtering to remove tissue fragments, the filtering is carried out for 2 times, and then the diluent is sucked and placed in a centrifuge tube for standby. The method comprises the following specific operations: and (4) continuing to perform the next male fish after finishing the whole artificial insemination process after dissecting 1 male fish.
Step 2, sperm motility detection:
and (3) immediately observing the sperm motility of the sperm diluent under an optical microscope, applying the semen with the sperm motility of more than 95 percent to the artificial insemination material, and ensuring the sperm motility of 4-tailed male fishes to be more than 95 percent.
Step 3, selecting female parent fish and performing artificial insemination:
selecting more than 74 sexually mature female parent fishes with age of more than 4 as experimental materials, anesthetizing the female parent fishes by using a special anesthetic MS-222 for live fishes with the concentration of 120-150 mg/L, placing the anesthetized female fishes on an operation table, carrying out artificial insemination operation by cooperation of two persons, covering the head of one person by using a wet towel and slightly fixing the head of the one person with force, then enabling the abdomen of the female fishes to be upward, enabling the fish bodies to be vertical to the operation table, enabling the other person to use a microsyringe (the scale is 1000 mu L) for experiments to absorb 150 mu L of sperm diluent, avoiding generating bubbles in the absorption and injection processes, slowly inserting the sperm diluent into a genital hole behind an anal orifice of the female fishes, and pressing the genital hole for 3-5 seconds by fingers after the injection is finished to prevent semen from flowing out. Putting the mixture into a culture pond for continuous culture after the completion;
step 4, controlling the breeding environment conditions after insemination:
the water temperature of the female parent fish after insemination is controlled to be above 9 ℃ in winter, the water temperature is gradually increased to 14 ℃ in 3-4 months, the cultivation temperature is prevented from greatly fluctuating, the salinity is 28-32, the pH is 7.8-8.2, the dissolved oxygen is more than or equal to 5mg/L, the running water is oxygenated and cultivated, and the like; feeding small yellow croaker, goby, shrimps, cuttlefish, clamworm, clam meat, scallop skirt and the like one month before farrowing for nutrition enhancement, adding vitamins and other mixing materials for feeding, and starting night illumination cultivation; stopping feeding for one week before farrowing; necessary breeding work such as feeding and excrement and residual bait removing is eliminated, illumination and artificial workshop movement are reduced, and a quiet environment is created for the breeding work;
step 5, counting the insemination success rate of the female parent fish in spring:
on 20 days 4 months in 2018, female parent fish start to breed successively, 68 total female parent fish with 74 fish can breed, the insemination success rate is 91.9 percent (insemination success rate = the number of tails of female parent fish capable of breeding fish/the number of tails of artificial insemination female fish x 100% = 68/74 x 100% = 91.9 percent), and the breeding number of each female fish is generally within the range of 10.2 x 104～25.5 × 104And (4) tail.
Example 2
The embodiment relates to an artificial insemination method of sebastes schlegeli hilgendorf fertilized fish in ying-style,
this example is substantially the same as example 1 except that the parent fish is of a different batch.
15-20 months 11 in 2018, the artificial insemination test of sebastes schlegeli is performed, 12 tails of anatomically mature male fishes are selected, and 120 tails of female parent fishes are artificially inseminated.
In 25 days 4 months in 2019, the female parent fish starts to breed successively, the number of the female parent fish capable of breeding is 112, the insemination success rate is 93.3 percent (insemination success rate = the number of tails of the female parent fish capable of breeding and the artificial insemination female fish tail number multiplied by 100% = 112/120 multiplied by 100% =93.3 percent), and the breeding number of each female fish ranges from 7.2 multiplied by 104～28.3 × 104And (4) tail.
Compared with the prior art, the invention has the following advantages:
(1) the invention relates to an artificial insemination method of sebastes schlegeli hilgendorf fertilized fish in ying and ying; an artificial insemination method of important oviparous economic fish sebastes schlegeli hilgendorf is established, the method replaces the direct collection of wild or cage-cultured brood parent fish for indoor artificial breeding, and solves the problems that the source of the brood parent fish is unstable, the transportation difficulty is high, the cost is increased, the transportation stress is easy to cause the abortion and death of the parent fish, the survival rate of the larval fish is low, the deformity rate is high and the like; the female parent fish with high fertility rate and high farrowing rate obtained by the method disclosed by the invention gets through the whole chain of full artificial breeding of sebastes schlegeli hilgendorf, fills the blank in the field, and provides technical conditions for large-scale breeding of offspring seeds and breeding.
(2) The invention relates to an artificial insemination method of sebastes schlegeli hilgendorf fertilized fish in ying and ying; by establishing an artificial insemination method of sebastes schlegeli hilgendorf, the insemination success rate of sexually mature female parent fish is more than 91 percent; the method has the advantages of feasible production practice, simple and easy operation, low cost and high efficiency, and can finish the fertilization work of even hundreds of female parent fishes by only using a few male fishes with sexual maturity.
(3) Compared with the indoor natural mating method, the artificial insemination method for the fertilized fish sebastes schlegeli hilgendorf in the ying body, disclosed by the invention, avoids the interference of culture conditions such as artificially controlled water temperature, illumination, water flow, sound and the like in the mating period, greatly improves the insemination success rate, further improves the fertilization rate and ensures the smooth breeding of the offspring seeds in the next year.
(4) According to the artificial insemination method for sebastes schlegeli hilgendorf fertilized fish in a nutrient body, the phenomenon of ' one male and more female ' and the phenomenon of one male and more female ' exist in the free mating of sebastes schlegeli hilgendorf, so that the trouble of family identification exists when a breeding family is constructed, and the method provides convenience for the construction of the family of sebastes schlegeli hildorf and the breeding of new lines and varieties.
The method has the significance that the insemination process of the female parent fish can be completed quickly by acquiring a small amount of sperms in the mating season of the male and female parent fish of sebastes schlegeli hilgendorf, the problem of low mating rate in the natural mating process of the indoor artificial breeding of the parent fish is solved, and the insemination success rate of the female parent fish is over 91 percent. The method of the invention thoroughly solves the defect of breeding by using wild or cage pregnant mares parent fish, can realize the full artificial breeding and the fry batch production of sebastes schlegeli hilgendorf, and also provides technical support for the breeding, breeding and proliferation releasing of sebastes schlegeli hilgendorf.
The foregoing description of specific embodiments of the present invention has been presented. It is to be understood that the present invention is not limited to the specific embodiments described above, and that various changes or modifications may be made by one skilled in the art within the scope of the appended claims without departing from the spirit of the invention.
Claims (10)
1. An artificial insemination method of fertilized fish sebastes schlegeli in ying, which is characterized by comprising the following steps:
step 1, preparing a sperm diluent and measuring sperm motility;
step 2, artificial insemination of female parent fish;
step 3, regulating and controlling the breeding environment after insemination;
and 4, counting the success rate of spring insemination.
2. The artificial insemination method of sebastes schlegeli hilgendorf fish as claimed in claim 1, wherein in step 1, the preparation of sperm diluent and the determination of sperm motility are specifically as follows:
dissecting the male parent fish to obtain a spermary, weighing and placing the spermary into a culture dish, and weighing the spermary: adding the diluent into the mixture according to the volume of 1:1-1: 1.5; shearing spermary with scissors or placing in a container to break the spermary, filtering tissue fragments with a filter screen for 2 times, sucking sperm diluent and placing in a centrifuge tube for use;
and (3) observing the sperm motility of the sperm diluent under an optical microscope, wherein the sperm diluent with the motility of more than 95 percent is used for artificial insemination materials for standby.
3. The artificial insemination method of sebastes schlegeli hilgendorf fish as claimed in claim 1, wherein in step 2, said artificial insemination of female parent fish is specifically: anaesthetizing female parent fish by using a live fish anesthetic MS-222 with the concentration of 120-150 mg/L, placing the live fish anesthetic MS-222 on an operation table, covering the head of a fish body by using a wet towel and fixing the wet towel slightly with force, enabling the belly of the female fish to face upwards to enable the fish body to be vertical to the operation table, sucking 150 mu L of sperm diluent by using a micro sample injector for experiments by using another person, slowly inserting the sperm diluent into a genital hole of the female fish, quickly injecting the sperm diluent into the fish body, and placing the parent fish into an experiment pool for continuous cultivation.
4. The artificial insemination method of sebastes schlegeli hilgendorf fish fertilized in ying-body as claimed in claim 1, wherein in step 3, the cultivation environment after insemination is specifically controlled as follows: controlling water temperature above 9 deg.C, starting night illumination cultivation and nutrition enhancement one month before farrowing, and stopping feeding one week before farrowing.
5. The artificial insemination method of sebastes schlegeli hilgendorf fish fertilized in ying-style according to claim 1, wherein in step 4, the statistics of fertilization farrowing rate are specifically as follows: and (3) measuring the insemination success rate of the female parent fish according to the situation of whether the parent fish farrows or not, continuously farrows the parent fish 4-5 months next year, recording the tails of the female parent fish which can farrow, and calculating the insemination success rate.
6. The artificial insemination method of sebastes schlegeli hilgendorf as a fish fertilized in ying body according to claim 1, wherein said male parent fish is a sexually mature male parent fish cultivated industrially at 3 ages or more, and is matured at 11 months per year in the testis.
7. The artificial insemination method of sebastes schlegeli hilgendorf as fish in vivo fertilization of claim 2, wherein said diluent is urine of male parent fish, and said urine is obtained by inserting a 2 mL syringe needle into bladder to suck urine when dissecting male parent fish, and storing said urine in a centrifuge tube for later use.
8. The artificial insemination method of sebastes schlegeli hilgendorf fish fertilized in camps as claimed in claim 6, wherein said sebastes schlegeli hilgendorf female parent fish is a sexually mature female of age 4 or more.
9. The artificial insemination method of sebastes schlegeli hilgendorf fish as claimed in claim 2, wherein said sperm diluent is sucked to avoid the generation of bubbles, and the germ cells are pressed by fingers for 3-5 s immediately after the injection to prevent the semen from flowing out.
10. The artificial insemination method of sebastes schlegeli hilgendorf fish of fertilization fish in vivo as claimed in claim 1, wherein said insemination success rate is calculated by using the formula of the tail of female parent fish capable of farrowing and the tail of artificial insemination female fish x 100%.
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN112931729A (en) * | 2021-03-31 | 2021-06-11 | 中国海洋大学 | Combined preparation for artificial induction parturition of oviparous schlegeli sebastes and application |
| CN115067322A (en) * | 2022-07-15 | 2022-09-20 | 中国海洋大学 | Method for high-activity non-freezing short-term preservation of sebastes schlegeli sperm |
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| CN103478036A (en) * | 2013-09-26 | 2014-01-01 | 宁波大学 | Cultivating method of marmoratus fish fries |
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| CN101658146A (en) * | 2008-08-25 | 2010-03-03 | 李思发 | Salt tolerant breeding technology for tilapia |
| JP2011067187A (en) * | 2009-09-25 | 2011-04-07 | Kagawa Prefecture | Artificial insemination method of internal fertilization fish of marine fish |
| CN103478036A (en) * | 2013-09-26 | 2014-01-01 | 宁波大学 | Cultivating method of marmoratus fish fries |
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| CN112931729A (en) * | 2021-03-31 | 2021-06-11 | 中国海洋大学 | Combined preparation for artificial induction parturition of oviparous schlegeli sebastes and application |
| CN115067322A (en) * | 2022-07-15 | 2022-09-20 | 中国海洋大学 | Method for high-activity non-freezing short-term preservation of sebastes schlegeli sperm |
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