CN106550909B - The method of fancy carp and megalobrame amblycephala subfamily distant hydridization - Google Patents
The method of fancy carp and megalobrame amblycephala subfamily distant hydridization Download PDFInfo
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- CN106550909B CN106550909B CN201610930546.6A CN201610930546A CN106550909B CN 106550909 B CN106550909 B CN 106550909B CN 201610930546 A CN201610930546 A CN 201610930546A CN 106550909 B CN106550909 B CN 106550909B
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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
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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
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Abstract
The invention discloses a kind of methods of fancy carp and megalobrame amblycephala subfamily distant hydridization, the following steps are included: being maternal parent population with fancy carp, megalobrama amblycephala is male parent parent population, artificial induced spawning is carried out to the maternal parent population and male parent parent population in mating period, dry method of artificial fertilization is carried out again, and obtained fertilized eggs are subjected to Lotic hatching in incubation tank, obtain the filial generation of fancy carp and megalobrama amblycephala, detection screening is carried out to the filial generation, obtains diploid fancy carp, diploid carp crucian carp and diploid crucian.The method of fancy carp and megalobrame amblycephala subfamily distant hydridization of the invention, not only incorporate the merit of fancy carp and megalobrama amblycephala, enrich market fish species, and good basis has been established for the breeding of excellent variety, valuable resource is provided for the breeding of subsequent novel Polyploid fish, there is important biological significance in biological evolution and genetics-breeding in fish.
Description
Technical field
The invention belongs to the distant hybridization fields between subfamily in fish hybridization field more particularly to cyprinid fish, specifically relate to
And a kind of method of fancy carp and megalobrame amblycephala subfamily distant hydridization.
Background technique
Hybridization of the affiliation more than inter-species or inter-species is commonly referred to as distant hybridization, is fish character improvement commonly side
Method.As a kind of important genetics-breeding in fish and character improvement method, it can integrate a whole set of foreign gene for distant hybridization, from
And change the expression regulation of filial generation gene, so that filial generation may be in the speed of growth, resistance, disease resistance and meat
Etc. show hybrid vigour.Filial generation as the inter-genera distant hybridization of red crucian carp and Xiangjiang wild carps is formed has figure excellent
Beauty, the speed of growth be fast, delicious meat, the advantages that premunition is strong, survival rate is high；The inter-subfamily distant hybridization shape of fancy carp and megalobrama amblycephala
At filial generation in addition to having the advantages that crucian carp carp hybridization fish or more, beautiful appearance, beautiful shape, in novel excellent class strain development
There is important value with genetic breeding aspect.
Existing biotechnology and method are how utilized and improved, suitable parent fish is selected in existing fish products kind and is carried out
Distant hybridization, to carry out genetic improvement to it, to obtain, shape is more excellent, kind is more abundant and more has to genetic research
The novel Polyploid fish of meaning just becomes those skilled in the art and faces for a long time and problem to be solved.
Compared with common carp, fancy carp is colorful, and scale and decorative pattern color are it is obvious that figure is smaller than common carp, fin
Item is more soft, has very high ornamental value, but vitality is markedly less than common carp, therefore purchasing price much compares carp
It is expensive.Carp is cultivated under natural conditions to be very easy to, and can be carried out breeding hatching when water temperature reaches 18 DEG C or more, and fancy carp is bred
Start a large amount of ovipositions generally after water temperature rises to 21 DEG C, egg-laying time is until from 5:00 AM or so to noon, during which if water
Temperature sharply declines suddenly, will interrupt oviposition, it is seen that it is very stringent to water temperature requirement to cultivate fancy carp.Fancy carp
(cyprinuscarpiohaematopterus) belong to Cypriniformes, the liploid fish (2n=100) of Cyprinidae, carp subfamily, megalobrama amblycephala
(Megalobramaamblycephala, 2n=48) belong to Cyprinidae, the liploid fish in Culter subfamily；In heredity, classification and figure
In terms of equal biological properties, all there is larger difference between fancy carp and megalobrama amblycephala, especially their chromosome numbers differ greatly,
Carp chromosome number is 100, and megalobrama amblycephala chromosome number is 48, is not compared between the two at multiple proportion and difference
Greatly, therefore distant hybridization is relatively difficult.So far domestic report between fancy carp and megalobrama amblycephala not yet.Explore a kind of fancy carp and group
The intermolecular hybrid technology of succeeding of head triangular bream has practical application value.
Summary of the invention
The technical problem to be solved by the present invention is to overcome the shortcomings of to mention in background above technology and defect, provide one
Kind rate of fertilization is high, and characters of progenies is excellent, clean-cut, remote between the fancy carp and megalobrama amblycephalas subfamily that be of great significance to genetic breeding
The method of edge hybridization.
In order to solve the above technical problems, technical solution proposed by the present invention are as follows:
A kind of method of fancy carp and megalobrame amblycephala subfamily distant hydridization, comprising the following steps: with fancy carp be maternal parent population, group
Head triangular bream is male parent parent population, carries out artificial induced spawning to the maternal parent population and male parent parent population in mating period, then to carry out dry method artificial
Obtained fertilized eggs are carried out in incubation tank Lotic hatching, obtain the filial generation of fancy carp and megalobrama amblycephala, to described miscellaneous by insemination
It hands over offspring to carry out detection screening, obtains diploid fancy carp, diploid carp crucian carp and diploid crucian.Incubation tank, which is that one kind is novel, incubates
Makeup is set, and having the advantage that 1) compared to Lentic hatching has endlessly fresh water source supplement, with air contact area
Greatly, the oxygen demand of fry hatching is greatly met；2) egg membrane ensure that hatching is good convenient for removing in time after fry hatching membrane
Water quality, while saving a large amount of labours；3) incubation tank is hatched and the seedling vigor being open is stronger, the seedling predation energy cultivated
Power is stronger.
Above-mentioned method, it is preferred that specifically includes the following steps:
(1) select within 3~4 months before breeding period that sexal maturity feature is obvious, the good female fancy carp of sign and male group head
Triangular bream carries out special Chi Peiyu respectively as the maternal parent population of distant hybridization and male parent parent population, is raised with smart bait within 1~2 month before breeding
It supports, it is primary every 1~3 day stimulation by running water；
(2) in the mid-May in the same year to early June, when water temperature stability is at 20 DEG C or more, for what is obtained after step (1)
The mixing ocyodinic of maternal parent population injection Luteinizing hormone releasing hormone analog and human chorionic gonadtropin is hastened parturition, 4~5h
The male parent parent population again to obtain after step (1) injects the mixing of Luteinizing hormone releasing hormone analog and human chorionic gonadtropin afterwards
Ocyodinic is hastened parturition, and maternal parent population and male parent parent population are put into same spawning pond by the quantity ratio of 1:6~8 after injection
In；
(3) in the maternal parent population and male parent parent population obtained after step (2), oviposition smoothly maternal parent population is selected, with production
Essence measures big male parent parent population and carries out artificial dry method insemination, obtained insemination ovum be placed in water temperature in 19 DEG C~20 DEG C of incubation tank into
Row Lotic hatching obtains the filial generation fry of fancy carp and megalobrama amblycephala；
(4) the filial generation fry obtained after step (3) is placed in incubation tank and continues culture 2~4 days, to the hybridization
Being transferred to after waist is selected occurs in offspring fry raised 1~3 day in the pond increased fertilizer in advance after splash soya-bean milk, to what is obtained after raising
Filial generation carries out detection screening, obtains diploid fancy carp, diploid carp crucian carp and diploid crucian.
In order to solve the problems, such as between fancy carp and megalobrama amblycephala that distant hybridization is difficult, the present invention by the way of artificial induced spawning,
Simultaneously in view of the vitality of fancy carp is weak higher compared with and to the sensibility of water temperature, the present invention selects Luteinizing hormone releasing hormone analog
With human chorionic gonadtropin as ocyodinic is mixed, to hasten parturition in suitable dosage injection parent population body, hatching rate is obvious
Increase, provides quantitative maximum possible for screening various Hybrid offsprings in this way；Then it is given and accepted using artificial dry method
Essence after 2~3 minute, the fertilized eggs for completing fertilization is equably arrived and are hatched into incubation tank.
Preferably, in the step (1), sign good behaviour is that figure is good, body colour is bright-coloured, physical strong and disease-free nothing
Wound；Using pectoral fin base portion without one needle injection method of abdominal cavity at squama when the injection.
Preferably, in the step (2), for the Luteinizing hormone releasing hormone analog and chorion of maternal parent population injection
In the mixing ocyodinic of promoting sexual gland hormone, the amount of Luteinizing hormone releasing hormone analog is 7~10ug/kg, and chorionic gonadotrophin swashs
The amount of element is 600~800IU/kg；For the Luteinizing hormone releasing hormone analog and chorionic gonadotrophin of the injection of male parent parent population
In the mixing ocyodinic of hormone, the amount of Luteinizing hormone releasing hormone analog is 5~6ug/kg, and the amount of human chorionic gonadtropin is
300~400IU/kg.Luteinizing hormone releasing hormone analog promotes parent population heat and oviposition, and human chorionic gonadtropin has rush
It into the double action of parent population ovulation and rush gonad development, is proved by a large number of experiments, above-mentioned injection dosage can make the production essence of parent population
Amount and egg laying amount reach maximum, and the injury simultaneously for parent population reaches minimum, and convenient for the control to the time, it is generally the case that
Carry out drug injection the dusk 18:00~20:00, and morning next day 8:00~9:00 can carry out fry production.
Preferably, in the step (2), the water surface area of spawning pond is 80cm2~90cm2。
In the step (3), the concrete operations of artificial dry method insemination include the following steps: for raun and milter to be put into same
A pond controls water temperature at 24~26 DEG C, is salvaged when milter knocks into the back raun, raun fish body is held in hand immediately first,
Left hand holds fish tail handle, and the right hand is held under the head of fish at ridge, and the body surface of fish obliquely, is gently dried, then gently by fish belly portion
Raun abdomen is pressed, ovum is flowed into dry ceramic bowl, the sperm of milter is extruded into above the ovum with same method, then
It is gently agitated for 2~3min with dry feather, is allowed to complete insemination.
Preferably, in the step (4), the method increased fertilizer in advance in pond specifically includes the following steps: be put into originally
Water exposure 1~2 day is put into well water and does not need to expose, and makes 70~80cm of the depth of water in pond, is then respectively adding 200~300ml
Green alga and 200~300ml cyanobacteria.
Preferably, in the step (4), the frequency for soya-bean milk of splashing is 1~2 times/day.
Preferably, the diploid crucian includes diploid red crucian, diploid flower crucian carp and the full white flower crucian carp of diploid.
Preferably, the detection screening includes shape measurement, the measurement of fluidic cell DNA content, renal tissue chromosome times
Property detection and blood cell shape identification.
Compared with prior art, the invention has the benefit that
1, the method for fancy carp of the invention and megalobrame amblycephala subfamily distant hydridization solves remote edge between fancy carp and megalobrama amblycephala
Hybridize difficult problem, the present invention considers that the vitality of fancy carp is weak compared with and to water temperature using artificial induced spawning by the way of
Sensibility is higher, and the present invention selects Luteinizing hormone releasing hormone analog to be used as with human chorionic gonadtropin and mixes ocyodinic, with
It hastens parturition in suitable dosage injection parent population body, hatching rate obviously increases, and mentions in this way for screening various Hybrid offsprings
Quantitative maximum possible has been supplied, the merit of fancy carp and megalobrama amblycephala is not only incorporated, has enriched market fish species, and
Good basis has been established for the breeding of excellent variety, has provided valuable resource for the breeding of subsequent novel Polyploid fish,
Biological evolution and genetics-breeding in fish have important biological significance.
2, the method for fancy carp of the invention and megalobrame amblycephala subfamily distant hydridization, obtained in offspring diploid fancy carp,
Diploid carp crucian carp and diploid crucian have also appeared gynogenesis fancy carp, and the speed of growth of these fishes is fast, and ornamental value is very
Height, the color such as diploid crucian is very bright-coloured, colorful, and figure is very beautiful, and apparent variation phenomenon occurs, especially
It is that the head of the flower crucian carp in diploid crucian type apparent punctation occurs, and this fish is very rare, in birds and flowers city
And beautiful scenery together with becoming in exhibition, be the characteristics of other distant hybridization at present do not have, market value is high.
And these fishes have extremely strong resistance (advantage of hybridization), can have certain warp as the parent of other cross combinations
Ji value, is worthy to be popularized.
3, fancy carp of the invention and megalobrame amblycephala subfamily distant hydridization provide a strong card for the origin of crucian
According to also to obtain red crucian carp on a large scale and crucian carp being spent to provide a kind of laboratory facilities, a parent for enriching other cross combinations comes
Source, the carp crucian carp that this cross combination obtains are a kind of offspring'ss (having mouth palpus on one side) for existing and obviously making a variation, illustrate that this remote edge is miscellaneous
It hands over to generate mutational variety and providing a reliable method；And the offspring that fancy carp and this cross combination of megalobrama amblycephala obtain, then
Being hybridized with other fishes, offspring's edible value of acquisition increases, such as the red crucian carp hybridized with fancy carp with megalobrama amblycephala,
Hybridize the offspring obtained with white crucian carp, advantageous will occur on edible value.
4, the incubation tank that the present invention uses is a kind of novel hatching apparatus, is had the advantage that compared to Lentic hatching
1) have endlessly that fresh water source supplement, large contact area with air greatly meet the oxygen demand of fry hatching；2) fry
Egg membrane ensure that the good water quality of hatching, while saving a large amount of labours convenient for removing in time after incubation of membrane；3) incubation tank is hatched
And the seedling vigor being open is stronger, the seedling prey ability cultivated is stronger.
Detailed description of the invention
In order to more clearly explain the embodiment of the invention or the technical proposal in the existing technology, to embodiment or will show below
There is attached drawing needed in technical description to be briefly described, it should be apparent that, the accompanying drawings in the following description is the present invention
Some embodiments for those of ordinary skill in the art without creative efforts, can also basis
These attached drawings obtain other attached drawings.
Fig. 1 is the outline drawing of diploid red crucian in fancy carp (♀) and megalobrama amblycephala (♂) filial generation.
Fig. 2 is the outline drawing of diploid flower crucian carp in fancy carp (♀) and megalobrama amblycephala (♂) filial generation.
Fig. 3 is the full white flower crucian carp outline drawing of diploid in fancy carp (♀) and megalobrama amblycephala (♂) filial generation.
Fig. 4 is diploid carp crucian carp outline drawing in fancy carp (♀) and megalobrama amblycephala (♂) filial generation.
Fig. 5 is diploid fancy carp outline drawing in fancy carp (♀) and megalobrama amblycephala (♂) filial generation.
Fig. 6 is the fluidic cell figure of diploid red crucian in fancy carp (♀) and megalobrama amblycephala (♂) filial generation.
Fig. 7 is the fluidic cell figure of diploid flower crucian carp in fancy carp (♀) and megalobrama amblycephala (♂) filial generation.
Fig. 8 is the fluidic cell figure of diploid carp crucian carp in fancy carp (♀) and megalobrama amblycephala (♂) filial generation.
Fig. 9 is diploid fancy carp fluidic cell figure in fancy carp (♀) and megalobrama amblycephala (♂) filial generation.
Figure 10 is diploid red crucian haemocyte figure in fancy carp (♀) and megalobrama amblycephala (♂) filial generation.
Figure 11 is diploid flower crucian carp crucian carp haemocyte figure in fancy carp (♀) and megalobrama amblycephala (♂) filial generation.
Figure 12 is diploid carp crucian carp haemocyte figure in fancy carp (♀) and megalobrama amblycephala (♂) filial generation.
Figure 13 is diploid fancy carp haemocyte figure in fancy carp (♀) and megalobrama amblycephala (♂) filial generation.
Figure 14 is diploid red crucian chromosome map in fancy carp (♀) and megalobrama amblycephala (♂) filial generation.
Figure 15 is diploid flower crucian carp chromosome map in fancy carp (♀) and megalobrama amblycephala (♂) filial generation.
Figure 16 is diploid carp crucian carp chromosome map in fancy carp (♀) and megalobrama amblycephala (♂) filial generation.
Figure 17 is diploid fancy carp chromosome map in fancy carp (♀) and megalobrama amblycephala (♂) filial generation.
Specific embodiment
To facilitate the understanding of the present invention, the present invention is done below in conjunction with Figure of description and preferred embodiment more complete
Face meticulously describes, but protection scope of the present invention is not limited to following specific embodiments.
Unless otherwise defined, all technical terms used hereinafter are generally understood meaning phase with those skilled in the art
Together.Technical term used herein is intended merely to the purpose of description specific embodiment, and it is of the invention to be not intended to limitation
Protection scope.
Unless otherwise specified, various raw material, reagent, the instrument and equipment etc. used in the present invention can pass through city
Field is commercially available or can be prepared by existing method.
Embodiment:
A kind of method of fancy carp and megalobrame amblycephala subfamily distant hydridization of the invention, comprising the following steps:
(1) 3~4 months before breeding period, select sexal maturity feature is obvious, sign is good, body colour is bright-coloured, it is physical strong and
Well-developed female fancy carp and male megalobrama amblycephala are respectively as the maternal parent population of distant hybridization and the special Chi Pei of male parent parent population progress
It educates, is raised with smart bait within 1~2 month before breeding, it is primary every 1~3 day stimulation by running water, to promote parent population gonadal maturation；
(2) in the mid-May in the same year to early June, when water temperature stability is at 20 DEG C or more, for what is obtained after step (1)
The mixing ocyodinic of maternal parent population injection Luteinizing hormone releasing hormone analog (LRH-A) and human chorionic gonadtropin (HCG) into
Row is hastened parturition, and the dosage of LRH-A is 7~10ug/kg, and the dosage of HCG is 600~800IU/kg；Again to obtain after step (1) after 4h
Male parent parent population injection Luteinizing hormone releasing hormone analog (LRH-A) and human chorionic gonadtropin (HCG) mixing ocyodinic
It hastens parturition, the dosage of LRH-A is 5~6ug/kg, and the dosage of HCG is 300~400IU/kg, by 1:6~8 after injection
It is 80cm that maternal parent population and male parent parent population are put into water surface area by quantity ratio2~90cm2Same spawning pond in, then regard regimen
It washes by water in time into pond, until the maternal parent population and male parent parent population start smoothly to lay eggs and produce essence；
(3) in the maternal parent population and male parent parent population obtained after step (2), oviposition smoothly maternal parent population is selected, with production
Essence measures big male parent parent population and is put into the same pond, controls water temperature at 24~26 DEG C, the case where observing the water surface is paid attention to, when milter chases after
It is salvaged when tail raun, raun fish body is held in hand immediately first, left hand holds fish tail handle, and the right hand is held in ridge under the head of fish
The body surface of fish obliquely (preferably with horizontal plane angle at 45 °), is gently dried by place, fish belly portion, then light pressure raun abdomen, makes ovum
Subflow enters in dry ceramic bowl, and the sperm of milter is extruded into above the ovum with same method, then light with dry feather
2~3min of light agitation, be allowed to complete insemination, and obtained insemination ovum pours into rapidly cut-and-dried water temperature and incubates for 19 DEG C~20 DEG C
Change in circuit and carry out Lotic hatching, obtains the filial generation fry of fancy carp and megalobrama amblycephala；
(4) the filial generation fry obtained after step (3) is placed in incubation tank and continues culture 2~4 days, to the hybridization
Raising occurs being transferred in the pond increased fertilizer in advance after waist point in offspring fry, soya-bean milk of splashing after 1~3 day, until fry grows to
It can normally ingest；Using shape measurement, the measurement of fluidic cell DNA content, the detection of renal tissue ploidy and haemocyte shape
State identification, carries out detection screening to the filial generation obtained after raising, obtains diploid fancy carp, diploid carp crucian carp and diploid crucian carp
Fish.
After filial generation obtained above is raised 3~4 months, the biological shapes phase of each filial generation of sampling Detection immediately
Index is closed, and is compared respectively with maternal fancy carp and male parent megalobrama amblycephala, the results are shown in Table 1:
Table 1: fancy carp, megalobrama amblycephala and the denumerable character of filial generation compare (unit: piece or item)
As shown in Table 1: five kinds of filial generation fishes are in shape (lateral line scales, Dorsal fin spine, pectoral fin item, abdomeinal fin item, stern fin ray)
Substantially endless to be congruent with maternal fancy carp and male parent megalobrama amblycephala (such as Fig. 1-5), illustrate the characteristics of filial generation fish existing forms make a variation.
On side line under squama and side line in terms of squama, the maternal character of filial generation all heredity.It is thin with streaming using common red crucian carp as reference
Born of the same parents' instrument measures red crucian carp and flower crucian carp, diploid carp crucian carp and diploid fancy carp (such as Fig. 6-9) in diploid crucian type, and DNA's is flat
Equal content is respectively 96.47,99.57,101.60 and 103.44, to illustrate that above-mentioned filial generation is all liploid fish.In order to
The morphological differences of identification of cell core, we are prepared for the blood film (such as Figure 10-13, scale 10um) of these four fishes, by comparing
It was found that diploid red crucian and diploid flower crucian carp haemocyte nucleus are in the same size, diploid carp crucian carp and diploid fancy carp haemocyte are thin
Karyon size is similar, more slightly smaller than diploid red crucian and diploid flower crucian carp nucleus.Again with renal tissue sectioning cells method to female
Property fancy carp and the filial generation chromosome number of somatic of male megalobrama amblycephala detected (such as Figure 14-17, scale 3um), pass through
The chromosome number for counting 5 kinds of hybridization fish offsprings of discovery is all 100, to prove that above-mentioned filial generation is all diploid.
The operating procedure of fluidic cell DNA content measurement are as follows: with the disposable syringe soaked through heparin from fish tail vein
Blood vessel blood sampling about 0.1mL, injection is equipped in the microcentrifugal tube (Eppendorf pipe) of 0.8% physiological saline, in blood and physiology
500 μ L nucleus extraction liquid DAPI-A(nuclei extraction solution, German Partec are added in salt water mixed liquid
Gmbh is provided), handle 10~12min of time；Sample is by 20 μm of nylon filters (German Partec GmbH is provided) filtering；DNA
Dyeing liquor (DAPI-B, German Partec GmbH are provided) is statically placed in dark place stained specimens about 10min, then upper machine testing.
The operating procedure of blood cell shape identification are as follows: 1) fish tail intravenous regional is wiped with iodine cotton swab, with disposable injection
Device takes 0.1ml(or takes 15 μ l using liquid-transfering gun) blood, touches a drop in clean slide one end, drop of blood is made to invest slide face
On；2) both ends that the glass slide is taken with a hand, hold rapidly one end of another glass slide with another hand, have blood in drop
Drop of blood is contacted on glass slide from the front to the back, makes two glass slides angle about at 45 °, gently moves, make drop of blood in line, then by preceding
It is a uniform thin slice to pusher；3) drying is placed on staining rack smear in air, and Wright's staining liquid is added dropwise, makes smear quilt
Dyeing liquor covering；4) it after dyeing 6 minutes, is uniformly slowly rinsed in the glass slide back side with distilled water, after seeing smear pinkiness, from
It so dries, last microscopy.
The operating procedure of renal tissue ploidy detection are as follows: right under 20 DEG C of water temperatures after cultural aim fish 2~3 days
It tests fish to inject PHA 1~3 time, each dosage is 2~8 μ g/g weight, and interval time is 12~24 hours, before dissection materials
2~6 hours, colchicine is injected, dosage is 2~4 μ g/g weight；Nephridial tissue is taken out, shreds nephridial tissue under physiological saline,
0.075mol/LKCL Hypotonic treatment, hypotonic 45 minutes or so at a temperature of 20 DEG C；With Ka Nuoshi fixer (glacial acetic acid: methanol=
1:3) fixed nephridial tissue 1~3 time；Piece is dripped on frost glass slide；Ji's nurse Sa (Giemsa) dye liquor dyes 55 minutes, thin water flow punching
Glass slide back side removal dye liquor is washed, rear microscopy is air-dried, takes pictures.
The method of the present invention obtains the hybridization fish of three types: and diploid crucian (diploid red crucian, diploid flower crucian carp, two
Times full white flower crucian carp of body), diploid carp crucian carp, diploid fancy carp.Above-mentioned filial generation color is very beautiful, and figure indicates very much, and
There is apparent variation phenomenon, for example the head of crucian carp is spent apparent punctation occur, there is half of mouth palpus in carp crucian carp, this
A little filial generation speeds of growth are fast, and ornamental value is high, possesses good market prospects.Fancy carp and megalobrama amblycephala subfamily of the invention
Between distant hybridization method, not only incorporate the merit of fancy carp and megalobrama amblycephala, enrich market fish species, and be excellent
Good basis has been established in the breeding of non-defective unit kind, valuable resource is provided for the breeding of subsequent novel Polyploid fish, in biology
It evolves and genetics-breeding in fish has important biological significance.
Claims (5)
1. a kind of method of fancy carp and megalobrame amblycephala subfamily distant hydridization, comprising the following steps:
(1) select within 3~4 months before breeding period that sexal maturity feature is obvious, the good female fancy carp of sign and male megalobrama amblycephala point
Special Chi Peiyu is not carried out as the maternal parent population of distant hybridization and male parent parent population, is raised with smart bait within 1~2 month before breeding, often
It is primary every 1~3 day stimulation by running water；
(2) in the mid-May in the same year to early June, when water temperature stability is at 20 DEG C or more, for the female parent obtained after step (1)
Parent population injection 7~10ug/kg of Luteinizing hormone releasing hormone analog and the mixing of 600~800IU/kg of human chorionic gonadtropin are urged
Agent is produced to hasten parturition, the male parent parent population after 4~5h again to obtain after step (1) inject Luteinizing hormone releasing hormone analog 5~
The mixing ocyodinic of 6ug/kg and 300~400IU/kg of human chorionic gonadtropin is hastened parturition, and 1:6~8 are pressed after injection
Quantity ratio maternal parent population and male parent parent population are put into same spawning pond；
(3) in the maternal parent population and male parent parent population obtained after step (2), oviposition smoothly maternal parent population is selected, is measured with essence is produced
Big male parent parent population is put into the same pond, controls water temperature at 24~26 DEG C, is salvaged immediately when milter knocks into the back raun, first
Raun fish body is held in hand, left hand holds fish tail handle, and the right hand is held under the head of fish at ridge, fish belly portion obliquely, gently
The body surface of fish is dried, then light pressure raun abdomen, flow into ovum in dry ceramic bowl, with same method by the sperm of milter
It is extruded into above the ovum, then is gently agitated for 2~3min with dry feather, be allowed to complete insemination, obtained insemination ovum is placed in
Lotic hatching is carried out in the incubation tank that water temperature is 19 DEG C~20 DEG C, obtains the filial generation fry of fancy carp and megalobrama amblycephala；
(4) the filial generation fry obtained after step (3) is placed in incubation tank and continues culture 2~4 days, to the filial generation
Being transferred to after waist is selected occurs in fry raised 1~3 day in the pond increased fertilizer in advance after splash soya-bean milk, the frequency for soya-bean milk of splashing is 1
~2 times/day, detection screening is carried out to the filial generation obtained after raising, obtains diploid fancy carp, diploid carp crucian carp and diploid
Crucian；
The method increased fertilizer in advance in pond specifically includes the following steps: being put into originally water exposure 1~2 day or being put into well water
It does not need to expose, makes 70~80cm of the depth of water in pond, be then respectively adding 200~300ml green alga and 200~300ml cyanobacteria.
2. the method for fancy carp according to claim 1 and megalobrame amblycephala subfamily distant hydridization, which is characterized in that the step
(1) in, sign good behaviour is that figure is good, body colour is bright-coloured, physical strong and disease-free and injury-free；Pectoral fin base portion is used when the injection
Without one needle injection method of abdominal cavity at squama.
3. the method for fancy carp according to claim 1 and megalobrame amblycephala subfamily distant hydridization, which is characterized in that the step
(2) in, the water surface area of spawning pond is 80cm2~90cm2。
4. the method for fancy carp described in any one of claim 1 to 3 and megalobrame amblycephala subfamily distant hydridization, feature exist
In the diploid crucian includes diploid red crucian, diploid flower crucian carp and the full white flower crucian carp of diploid.
5. the method for fancy carp described in any one of claim 1 to 3 and megalobrame amblycephala subfamily distant hydridization, feature exist
In the detection screening includes that shape measurement, the measurement of fluidic cell DNA content, the detection of renal tissue ploidy and blood are thin
Born of the same parents' Morphological Identification.
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| CN109329128B (en) * | 2018-10-31 | 2022-05-17 | 湖南师范大学 | Method for establishing Japanese white crucian carp and red crucian carp hybrid strain and method for cultivating cross crucian carp No. 2 |
| CN110269010A (en) * | 2019-07-29 | 2019-09-24 | 天津农学院 | The method of oriental cherry gold-plating goldfish germplasm initiative |
| CN110915727B (en) * | 2019-12-10 | 2021-02-12 | 湖南师范大学 | Method for breeding multiple colorful double-tail goldfish through distant hybridization |
| CN115735858B (en) * | 2022-11-24 | 2024-04-19 | 中国科学院水生生物研究所 | Method for efficiently creating new polyploid crucian by sterility synthesis |
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