CN107155897A - A kind of muskmelon flower pesticide inducing culture and Anther culture breeding method - Google Patents
A kind of muskmelon flower pesticide inducing culture and Anther culture breeding method Download PDFInfo
- Publication number
- CN107155897A CN107155897A CN201710589435.8A CN201710589435A CN107155897A CN 107155897 A CN107155897 A CN 107155897A CN 201710589435 A CN201710589435 A CN 201710589435A CN 107155897 A CN107155897 A CN 107155897A
- Authority
- CN
- China
- Prior art keywords
- culture
- muskmelon
- flower pesticide
- bud
- anther
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Classifications
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H4/00—Plant reproduction by tissue culture techniques ; Tissue culture techniques therefor
- A01H4/005—Methods for micropropagation; Vegetative plant propagation using cell or tissue culture techniques
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H4/00—Plant reproduction by tissue culture techniques ; Tissue culture techniques therefor
- A01H4/001—Culture apparatus for tissue culture
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Biotechnology (AREA)
- Developmental Biology & Embryology (AREA)
- Cell Biology (AREA)
- Botany (AREA)
- Environmental Sciences (AREA)
- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
Abstract
The invention discloses a kind of muskmelon flower pesticide inducing culture and Anther culture breeding method, belong to fruits and vegetables and cultivate planting technology field.The muskmelon flower pesticide inducing culture that the present invention is researched and developed contains a great number of elements, trace element, molysite, organic principle, inorganic additive, plant growth regulator, coagulator and other additives etc..The muskmelon Anther culture breeding method and step of the present invention includes：Bud materials and pretreatment, the preparation of culture medium, flower pesticide inoculation, flower pesticide Fiber differentiation, flower pesticide differentiation culture, domestication and transplanting etc..Fiber differentiation is carried out to muskmelon flower pesticide using the present invention, antherderived callus inductivity can be effectively improved, muskmelon Anther culture breeding process is greatly accelerated.
Description
Technical field
The present invention relates to a kind of muskmelon flower pesticide inducing culture and Anther culture breeding method, and in particular to one kind effectively improves flower
Medicine Callus induction rate, the muskmelon flower pesticide inducing culture and Anther culture breeding method for greatly accelerating muskmelon Anther culture breeding process, category
Planting technology field is cultivated in fruits and vegetables.
Background technology
Muskmelon（melon）, the sweet melon of alias, muskmelon, intestines melon, fruit melon and ripe melon are worn, cucurbit are belonged in Plant Taxonomy
Section（Cucurbitaceae）Cucumis（Cucumis）, it is 1 year raw herbaceous plant, is a kind of fruit being loved by people
Type vegetables.Muskmelon originates in India, the tropical desert area in Africa, bright about in period in the Northern Wei Dynasty as watermelon together passes to China
Towards beginning plantation extensively.It is famous with its unique flavor always as one of big fruit in the world ten.
Contain substantial amounts of carbohydrate and citric acid, carrotene and vitamin B, C etc., and water in ripe melon fruit
Divide abundant, can relieve summer heat heat-clearing, quench one's thirst of promoting the production of body fluid, relieving restlessness etc.；Contain invertase in nutrition muskmelon, can be by insoluble protein qualitative change
Into soluble protein, kidney patient can be helped to absorb nutrition, it is beneficial to nephrotic；Cucurbitacin B contained by muskmelon pedicel can be bright
The aobvious experimental liver glycogen accumulation of increase, mitigates chronic liver injury, so as to prevent hepatic cell fattydegeneration and suppress fibroplasia；Sweet tea
Muskmelon pedicel contains picrotoxin, the crystallinity amaroid such as Cucurbitacin B, E, can stimulate gastric mucosa, takes orally appropriate, can cause vomiting, but be not
Body absorbs, and without the drawback such as collapse and poisoning；Modern study finds that muskmelon seed kills the effect such as roundworm, filaria, can be extensive
For treating malnutrition due to parasitic infestation illness.Extra-nutrition muskmelon cold cuts contain protein, fat, carbohydrate, inorganic salts etc., can supplement people
Energy and nutrient required for body, help body recovery health.
As a kind of important melon industrial crops, muskmelon is one of big important fruit in the world ten, cultivated area and yield
Larger, world temperate zone to torrid areas is cultivated extensively.The states such as China, Russia, Spain, the U.S., Iran, Italy, Japan
Family generally cultivates, with Chinese yield highest.Since 1980s, the cultivated area and yield of China's muskmelon are occupy always
The first in the world.Therefore, China's muskmelon production occupies critical role in the world, and muskmelon is the high-end agriculture dominant crop of China,
To increasing peasant income and agricultural restructuring important in inhibiting.
Because muskmelon is cross-pollinatd plant, natural hybrization rate is high, therefore is had using conventional breeding means acquisition pure lines
The shortcomings of cycle length, difficulty are big, inhereditary feature is unstable, and these problems can be just avoided using monoploid technology Breeding, greatly
It is big to improve breeding efficiency.Haploid breeding technology refers to make the heterogeneous gamete of some kinds develop into list by inducing parthenogenesis
Times body plant, then be sheerly by chromosome doubling, afterwards a kind of breeding method of progress breed breeding.This technology can
Shorten the breeding time limit, improve breeding efficiency.But in nature, the probability of the spontaneous Haploid production plant of crop is extremely low, Bu Guotong
Monoploid, such as flower pesticide or isolated microspore culture technique can be obtained by crossing some artificial means.So far, do not found in muskmelon
The haplobiont of spontaneous generation, so muskmelon flower training technology just turns into the important channel for obtaining muskmelon haplobiont.
The content of the invention
The purpose of the present invention is lacked for current muskmelon flower training callus induction rate is extremely low, flower training technology is incomplete
It is sunken that there is provided a kind of muskmelon flower pesticide induction for effectively improving antherderived callus inductivity, greatly accelerating muskmelon Anther culture breeding process
Culture medium and Anther culture breeding method.
The technical solution adopted by the present invention is as follows：
A kind of muskmelon flower pesticide inducing culture, it is characterised in that：Culture medium prescription following components content point in every liter of distilled water
It is not：KNO380-90mg/L, MgSO4∙7H2O 140-160mg/L, KH2PO470-76mg/L, Ca (NO3)2∙4H2O 160-
200mg/L, MnSO4∙4H2O 25-30mg/L, Na2- EDTA 11-14mg/L, FeSO4∙7H2O 12-15mg/L, H3BO3
8.0-8.5mg/L, ZnSO4∙7H2O 8.5-9.5mg/L, KI 0.45-0.55mg/L, AgNO315-19mg/L, Na2MoO4∙
2H2O 0.2-0.3mg/L, CuSO4∙5H2O 0.02-0.03mg/L, CoCl2∙6H2O 0.02-0.03mg/L, adenine 13-
17mg/L, glutamine 450-550mg/L, vitamin B1 0.9-1.1mg/L, vitamin B6 0.6-0.8mg/L, Vc
0.55-0.65mg/L, nicotinic acid 4.5-5.5mg/L, inositol 120-140mg/L, glycine 3.5-4.5mg/L, insulin
1.8-2.2mg/L, cyclic guanylic acid 0.15-0.25g/L, glutathione 30-35mg/L, ethylmethane sulfonate 1.0-1.4g/L,
Sucrose 42-48g/L, agar 6.5-7.5g/L, 6-BA 0.8-1.2mg/L, NAA 0.8-1.2mg/L, mannitol 14-16g/
L, coconut palm breast 27-33g/L, plant vulcanization kinetin PSK- α 18-22pmol/L.
Described culture medium prescription following components optimum content in every liter of distilled water is respectively：KNO385mg/L, MgSO4
∙7H2O 150mg/L, KH2PO473mg/L, Ca (NO3)2∙4H2O 180mg/L, MnSO4∙4H2O 27.5mg/L, Na2-EDTA
12.5mg/L, FeSO4∙7H2O 13.5mg/L, H3BO38.25mg/L, ZnSO4∙7H2O 9.0mg/L, KI 0.5mg/L,
AgNO317mg/L, Na2MoO4∙2H2O 0.25mg/L, CuSO4∙5H2O 0.025mg/L, CoCl2∙6H2O 0.025mg/L, gland
Purine 15mg/L, glutamine 500mg/L, vitamin B1 1.0mg/L, vitamin B6 0.7mg/L, Vc 0.6mg/L, nicotinic acid
5.0mg/L, inositol 130mg/L, glycine 4.0mg/L, insulin 2.0mg/L, cyclic guanylic acid 0.2g/L, glutathione
32.5mg/L, ethylmethane sulfonate 1.2g/L, sucrose 45g/L, agar 7.0g/L, 6-BA 1.0mg/L, NAA 1.0mg/L,
Mannitol 15g/L, coconut palm breast 30g/L, plant vulcanization kinetin PSK- α 20pmol/L.
The adding method of described ethylmethane sulfonate is：Prepare after ethylmethane sulfonate solution, with 0.22 μ of sterilizing
M filtering with microporous membrane sterilizing, when condensing to 45-55 DEG C after culture medium high pressure steam sterilization, taking-up is placed in superclean bench
In, sterile working adds ethylmethane sulfonate solution, shakes up in condensation, 24h times and is inoculated with.
The secure ph of described culture medium is：5.4-5.8, optimal secure ph is 5.6.
A kind of muskmelon Anther culture breeding method, it is characterised in that：Anther culture breeding method is operated as follows：
（1）Bud is drawn materials and pretreatment
When in crop field, the muskmelon of conventional cultivation is in initial bloom stage, in morning 9-11 points collection robust growth, the plant of no disease and pests harm
It is examination material to take developmental stage to be in the bud of mid-late uninucleate stage, and self-sealing plastic bag is used again after the bud of collection is wrapped up with wet gauze
Cover, be placed in low-temperature treatment 2d in 4 DEG C of refrigerators, be then transferred to Heat thermostability 2d under 36 DEG C of environment；
（2）The preparation of culture medium
It is sub-packed in after culture medium described in claim 1,2,3 and 4 is prepared in 250ml triangular flask, every bottle of Sheng 50ml-
Be placed in after 70ml culture medium, sealing temperature be 121 DEG C, pressure be the 20min that sterilizes under the conditions of 15kPa high steams, condense to
At 45-55 DEG C, taking-up is placed in superclean bench, sterile working, is added ethylmethane sulfonate solution, is shaken up condensation, 24h times
Interior inoculation；
（3）Flower pesticide is inoculated with
Pretreated bud is taken out, first 1-2 min is rinsed with flowing water, peels off anthocaulus with tweezers, use fritter gauze wrapped, is immersed
15s in 70% alcohol, is sterilized 8-10 minutes with being transferred to after aseptic water washing 2-3 times in 0.1% mercuric chloride solution, is then rushed with sterilized water
Wash 3-4 times, bud is put into the sterilizing culture dish for being covered with filter paper, flower pesticide is stripped under aseptic condition be inoculated into step 2 and prepare
Inducing culture on, every bottle access 14-16 pieces of flower pesticide；
（4）Flower pesticide Fiber differentiation
By be inoculated with flower pesticide after culture medium be placed in temperature for 26-28 DEG C, intensity of illumination 1400-1800lx, photoperiod 11h light/13h
Cultivated under dark environment, induce antherderived callus；
（5）Flower pesticide differentiation culture
After after antherderived callus length to diameter 2.5cm, select translucent, loosely organized callus and transfer on differential medium
Differentiation culture is carried out, callus gradually differentiates green seedling；
（6）Domestication is with transplanting
Green seedling growth is vigorous after flower pesticide differentiation culture 35d, takes out test tube seedling after sealed membrane now is opened into domestication hardening, 7d,
Root is cleaned up, greenhouse, conventional cultivation is transplanted into.
The step（1）The muskmelon bud selection standard of mid-late uninucleate stage be：The vertical stem length degree of muskmelon bud is measured, is indulged
Stem length degree is 3.0-3.6mm bud.
The step（5）Breaking up the condition cultivated is：25-27 DEG C of temperature, humidity are 70%-80%, intensity of illumination 2000-
2500lux, photoperiod 12h light/12h are dark.
The muskmelon flower pesticide inducing culture and Anther culture breeding method that the present invention is provided are the research in existing other vegetable seeds
Specificity requires to carry out the achievement of creative improvement for muskmelon Anther Culture on basis, and the present invention substantially increases muskmelon flower
Medicine callus induction rate, establishes muskmelon Anther culture breeding program first.
Muskmelon flower pesticide inducing culture provided by the present invention, is screened by a large amount of single-factors, multifactor experiment,
The nutrient media components proportioning that the present invention is demonstrated by substantial amounts of experimental study is optimum combination.Using training provided by the present invention
Foster base can greatly improve muskmelon induction of anther callus efficiency, therefore with higher industry promotional value.
Embodiment
With reference to embodiment, the present invention is further illustrated.
A kind of muskmelon flower pesticide inducing culture and Anther culture breeding method, it is characterised in that：Anther culture breeding method is by following step
Rapid operation：
（1）Bud is drawn materials and pretreatment
When in crop field, the muskmelon of conventional cultivation is in initial bloom stage, in morning 9-11 points collection robust growth, the plant of no disease and pests harm
It is examination material to take developmental stage to be in the bud of mid-late uninucleate stage, and the muskmelon bud selection standard of mid-late uninucleate stage is：Measure muskmelon
The vertical stem length degree of bud, indulges the bud that stem length degree is 3.0-3.6mm；Again with self-styled modeling after the bud of collection is wrapped up with wet gauze
Pocket covers, and is placed in low-temperature treatment 2d in 4 DEG C of refrigerators, is then transferred to Heat thermostability 2d under 36 DEG C of environment；
（2）The preparation of culture medium
Muskmelon flower pesticide inducing culture is prepared, culture medium prescription following components content in every liter of distilled water is respectively：KNO3
80-90mg/L, MgSO4∙7H2O 140-160mg/L, KH2PO470-76mg/L, Ca (NO3)2∙4H2O 160-200mg/L,
MnSO4∙4H2O 25-30mg/L, Na2- EDTA 11-14mg/L, FeSO4∙7H2O 12-15mg/L, H3BO3 8.0-8.5mg/
L, ZnSO4∙7H2O 8.5-9.5mg/L, KI 0.45-0.55mg/L, AgNO315-19mg/L, Na2MoO4∙2H2O 0.2-
0.3mg/L, CuSO4∙5H2O 0.02-0.03mg/L, CoCl2∙6H2O 0.02-0.03mg/L, adenine 13-17mg/L, paddy ammonia
Acid amides 450-550mg/L, vitamin B1 0.9-1.1mg/L, vitamin B6 0.6-0.8mg/L, Vc 0.55-0.65mg/L,
Nicotinic acid 4.5-5.5mg/L, inositol 120-140mg/L, glycine 3.5-4.5mg/L, insulin 1.8-2.2mg/L, ring bird
Thuja acid 0.15-0.25g/L, glutathione 30-35mg/L, ethylmethane sulfonate 1.0-1.4g/L, sucrose 42-48g/L, fine jade
Fat 6.5-7.5g/L, 6-BA 0.8-1.2mg/L, NAA 0.8-1.2mg/L, mannitol 14-16g/L, coconut palm breast 27-33g/L,
Plant vulcanizes kinetin PSK- α 18-22pmol/L, and pH value is：5.4-5.8；
Culture medium prescription following components optimum content in every liter of distilled water is respectively：KNO385mg/L, MgSO4∙7H2O
150mg/L, KH2PO473mg/L, Ca (NO3)2∙4H2O 180mg/L, MnSO4∙4H2O 27.5mg/L, Na2-EDTA 12.5mg/
L, FeSO4∙7H2O 13.5mg/L, H3BO38.25mg/L, ZnSO4∙7H2O 9.0mg/L, KI 0.5mg/L, AgNO3 17mg/
L, Na2MoO4∙2H2O 0.25mg/L, CuSO4∙5H2O 0.025mg/L, CoCl2∙6H2O 0.025mg/L, adenine 15mg/L,
Glutamine 500mg/L, vitamin B1 1.0mg/L, vitamin B6 0.7mg/L, Vc 0.6mg/L, nicotinic acid 5.0mg/L, flesh
Alcohol 130mg/L, glycine 4.0mg/L, insulin 2.0mg/L, cyclic guanylic acid 0.2g/L, glutathione 32.5mg/L, first
Base sulfonic acid 1.2g/L, sucrose 45g/L, agar 7.0g/L, 6-BA 1.0mg/L, NAA 1.0mg/L, mannitol 15g/
L, coconut palm breast 30g/L, plant vulcanization kinetin PSK- α 20pmol/L, optimal secure ph is 5.6；
It is sub-packed in after culture medium is prepared in 250ml triangular flask, every bottle of Sheng 50ml-70ml culture medium is placed in after sealing
Temperature is 121 DEG C, pressure is the 20min that sterilizes under the conditions of 15kPa high steams, and when condensing to 45-55 DEG C, taking-up is placed in ultra-clean work
Make in platform, sterile working, add ethylmethane sulfonate solution, the adding method of ethylmethane sulfonate is：Prepare pyrovinic acid
After ethyl ester solution, sterilized with 0.22 μm of filtering with microporous membrane of sterilizing, after condensing to 45-55 after culture medium high pressure steam sterilization
DEG C when, taking-up is placed in superclean bench, sterile working, is added ethylmethane sulfonate solution, is shaken up condensation, and the 24h times are inscribed
Kind；Shake up in condensation, 24h times and be inoculated with；
（3）Flower pesticide is inoculated with
Pretreated bud is taken out, first 1-2 min is rinsed with flowing water, peels off anthocaulus with tweezers, use fritter gauze wrapped, is immersed
15s in 70% alcohol, is sterilized 8-10 minutes with being transferred to after aseptic water washing 2-3 times in 0.1% mercuric chloride solution, is then rushed with sterilized water
Wash 3-4 times, bud is put into the sterilizing culture dish for being covered with filter paper, flower pesticide is stripped under aseptic condition be inoculated into step 2 and prepare
Inducing culture on, every bottle access 14-16 pieces of flower pesticide；
（4）Flower pesticide Fiber differentiation
By be inoculated with flower pesticide after culture medium be placed in temperature for 26-28 DEG C, intensity of illumination 1400-1800lx, photoperiod 11h light/13h
Cultivated under dark environment, induce antherderived callus；
（5）Flower pesticide differentiation culture
After after antherderived callus length to diameter 2.5cm, select translucent, loosely organized callus and transfer on differential medium
Differentiation culture is carried out, the condition of differentiation culture is：25-27 DEG C of temperature, humidity be 70%-80%, intensity of illumination 2000-2500lux,
Photoperiod 12h light/12h is dark, and callus gradually differentiates green seedling；
（6）Domestication is with transplanting
Green seedling growth is vigorous after flower pesticide differentiation culture 35d, takes out test tube seedling after sealed membrane now is opened into domestication hardening, 7d,
Root is cleaned up, greenhouse, conventional cultivation is transplanted into.
Embodiment 1
A kind of muskmelon flower pesticide inducing culture, is prepared according to formula as below：
Following components content is respectively in every liter of distilled water：KNO385mg/L, MgSO4∙7H2O 150mg/L, KH2PO4 73mg/
L, Ca (NO3)2∙4H2O 180mg/L, MnSO4∙4H2O 27.5mg/L, Na2- EDTA 12.5mg/L, FeSO4∙7H2O 13.5mg/
L, H3BO38.25mg/L, ZnSO4∙7H2O 9.0mg/L, KI 0.5mg/L, AgNO317mg/L, Na2MoO4∙2H2O
0.25mg/L, CuSO4∙5H2O 0.025mg/L, CoCl2∙6H2O 0.025mg/L, adenine 15mg/L, glutamine
500mg/L, vitamin B1 1.0mg/L, vitamin B6 0.7mg/L, Vc 0.6mg/L, nicotinic acid 5.0mg/L, inositol 130mg/
L, glycine 4.0mg/L, insulin 2.0mg/L, cyclic guanylic acid 0.2g/L, glutathione 32.5mg/L, pyrovinic acid second
Ester 1.2g/L, sucrose 45g/L, agar 7.0g/L, 6-BA 1.0mg/L, NAA 1.0mg/L, mannitol 15g/L, coconut palm breast
30g/L, plant vulcanization kinetin PSK- α 20pmol/L, optimal secure ph is 5.6.
It is sub-packed in after culture medium is prepared in 250ml triangular flask, every bottle of Sheng 50ml-70ml culture medium, after sealing
Be placed in temperature be 121 DEG C, pressure be the 20min that sterilizes under the conditions of 15kPa high steams, when condensing to 45-55 DEG C, taking-up is placed in super
In net workbench, sterile working adds ethylmethane sulfonate solution, and the adding method of ethylmethane sulfonate is：Prepare methyl
After sulfonic acid solution, sterilized, condensed to after after culture medium high pressure steam sterilization with 0.22 μm of filtering with microporous membrane of sterilizing
At 45-55 DEG C, taking-up is placed in superclean bench, sterile working, is added ethylmethane sulfonate solution, is shaken up condensation, 24h times
Interior inoculation；Shake up in condensation, 24h times and be inoculated with.
Experiment melon variety elects red city 10 and imperial sweet tea one as, and Anther culture breeding method is operated as follows：
Bud is drawn materials and pretreatment：When in crop field, the muskmelon of conventional cultivation is in initial bloom stage, morning 9-11 points collection robust growth,
It is examination material, the muskmelon bud selection of mid-late uninucleate stage to take developmental stage to be in the bud of mid-late uninucleate stage on the plant of no disease and pests harm
Standard is：The vertical stem length degree of muskmelon bud is measured, the bud that stem length degree is 3.0-3.6mm is indulged；By the bud wet gauze of collection
Covered again with self-sealing plastic bag after parcel, be placed in low-temperature treatment 2d in 4 DEG C of refrigerators, be then transferred to Heat thermostability under 36 DEG C of environment
2d；
Flower pesticide is inoculated with：Pretreated bud is taken out, first 1-2 min is rinsed with flowing water, peels off anthocaulus with tweezers, use fritter gauze
15s in parcel, 70% alcohol of immersion, with sterilization 9 minutes in 0.1% mercuric chloride solution are transferred to after aseptic water washing 2-3 time, then with nothing
Bacterium water is rinsed 3-4 times, bud is put into the sterilizing culture dish for being covered with filter paper, stripping flower pesticide under aseptic condition is inoculated into step 2
On the inducing culture prepared, every bottle of 14-16 pieces of flower pesticide of access；
Flower pesticide Fiber differentiation：By be inoculated with flower pesticide after culture medium be placed in temperature for 27 DEG C, intensity of illumination 1600lx, photoperiod 11h
Cultivated under environment dark light/13h, induce antherderived callus, count induction of anther callus rate, anther callus is lured
Conductance（%）=anther callus number/inoculation flower pesticide number × 100%, actual count obtains red city 10 and is cured with imperial number flower pesticide of sweet tea
Injured tissue inductivity is respectively 35.4% and 28.8%；
Flower pesticide differentiation culture：Transferred after after antherderived callus length to diameter 2.5cm, selecting translucent, loosely organized callus
Enter progress differentiation culture on differential medium, the condition of differentiation culture is：26 DEG C of temperature, humidity are 75%, intensity of illumination
2250lux, photoperiod 12h light/12h are dark；Callus gradually differentiates green seedling；
Domestication is with transplanting：Green seedling growth is vigorous after flower pesticide differentiation culture 35d, and now opening sealed membrane will after domestication hardening, 7d
Test tube seedling is taken out, and root is cleaned up, greenhouse, conventional cultivation is transplanted into.
Embodiment 2
In the R＆D process of the muskmelon flower pesticide Fiber differentiation of the present invention, the single-factor proportioning test of nutrient media components has been carried out,
The single factor experiment constituent species of culture medium prescription are respectively：Adenine, Vc, insulin, cyclic guanylic acid, ethylmethane sulfonate,
Coconut palm breast and plant vulcanization kinetin PSK- α.Each experiment compositional difference only limits one kind, and other components are same as Example 1.Examination
Test melon variety and also elect red city 10 and imperial sweet tea one as, the compound method of culture medium, Anther culture breeding method with embodiment 1
It is identical.
Single-factor proportioning test is specifically matched and result of the test such as following table 1- tables 7.
The single factor experiment changed by the muskmelon flower pesticide inducing culture component of table 1 to table 7 is it can be found that culture medium prescription
Single factor experiment constituent species in insulin, the increase of ethylmethane sulfonate and coconut palm breast proportioning generate significant unfavorable effect
Should；And the increase that adenine, Vc, cyclic guanylic acid and plant vulcanize kinetin PSK- α can not dramatically increase the induction effect of culture medium
Really, because the increase of these additives can improve the preparation cost of culture medium, the minimum of optimal value is reached in culture medium inducing effect
Proportional quantity is the optimum proportioning after comprehensive production cost.As can be seen here, carrying out any increasing or decreasing for nutrient media components is
Unfavorable, while illustrating that the component proportion of the present invention is optimal.
Embodiment 3
For muskmelon flower pesticide inducing culture relatively more provided by the present invention with other culture mediums to muskmelon flower pesticide embryo callus subculture group
The difference of induced efficiency is knitted, we have selected 2 kinds of flower pesticide inducing cultures in addition, experiment kind is also using red city 10 and imperial sweet tea
No. one, flower pesticide separation method, tissue culture operating method, cultural method are same as Example 1, and embryo callus is distinguished after being formed
Count induction of anther callus rate of the red city 10 with the flower pesticide of imperial sweet tea one in 2 kinds of culture mediums.
Other 2 kinds of flower pesticide Fiber differentiation based formulas are：
C17-2 culture mediums：C17-2 minimal medium+20g/L sucrose+7g/L agar+2mg/L 2,4-D+1mg/L kinetins, pH
For 5.8（Documents come from F S Wen etc.《Sorghum flower pesticide and inflorescence culture evoked callus and regeneration plant》）；
MS culture mediums：Sucrose+1.0mg/LNAA+1.0mg/L KT+10% the coconut palms of MS minimal mediums+3% breast, pH is 5.8（It is right
Come from high elegant cloud etc. than file《Plantlets of Tomato Obtained From Anther Culture In Vitro》）.
Culture medium contrast experiment
The other 2 kinds of culture mediums induction provided using flower pesticide inducing culture provided by the present invention and comparative example is red
Induction of anther callus rate obtained by number anther callus statistics in city 10 and imperial sweet tea is compared, and comparative result is as follows
Shown in table 8.
By upper table 8 it can be found that muskmelon flower pesticide inducing culture and Anther culture breeding method that the present invention is provided are to muskmelon product
The induction of anther callus rate average of kind red city 10 and imperial sweet tea one is up to 32.1%, it can be found that the muskmelon flower of the present invention
Medicine inducing culture and Anther culture breeding method specificity require the achievement of the creative improvement of progress for muskmelon Anther Culture, first
Establish muskmelon Anther culture breeding program.Muskmelon anther callus can be greatly improved using culture medium provided by the present invention to lure
Efficiency is led, therefore with higher industry promotional value.
Those skilled in the art can be according to present disclosure and the art technology grasped in the present invention
Appearance makes replacement or modification, but these are replaced or modification is all not regarded as a departure from present inventive concept, and these are replaced or modification
In claimed interest field.
Claims (7)
1. a kind of muskmelon flower pesticide inducing culture, it is characterised in that：Culture medium prescription following components content in every liter of distilled water
Respectively：KNO380-90mg/L, MgSO4∙7H2O 140-160mg/L, KH2PO470-76mg/L, Ca (NO3)2∙4H2O 160-
200mg/L, MnSO4∙4H2O 25-30mg/L, Na2- EDTA 11-14mg/L, FeSO4∙7H2O 12-15mg/L, H3BO3
8.0-8.5mg/L, ZnSO4∙7H2O 8.5-9.5mg/L, KI 0.45-0.55mg/L, AgNO315-19mg/L, Na2MoO4∙
2H2O 0.2-0.3mg/L, CuSO4∙5H2O 0.02-0.03mg/L, CoCl2∙6H2O 0.02-0.03mg/L, adenine 13-
17mg/L, glutamine 450-550mg/L, vitamin B1 0.9-1.1mg/L, vitamin B6 0.6-0.8mg/L, Vc
0.55-0.65mg/L, nicotinic acid 4.5-5.5mg/L, inositol 120-140mg/L, glycine 3.5-4.5mg/L, insulin
1.8-2.2mg/L, cyclic guanylic acid 0.15-0.25g/L, glutathione 30-35mg/L, ethylmethane sulfonate 1.0-1.4g/L,
Sucrose 42-48g/L, agar 6.5-7.5g/L, 6-BA 0.8-1.2mg/L, NAA 0.8-1.2mg/L, mannitol 14-16g/
L, coconut palm breast 27-33g/L, plant vulcanization kinetin PSK- α 18-22pmol/L.
2. the muskmelon flower pesticide inducing culture according to claim 1, it is characterised in that：Described culture medium prescription is by every
Rising following components optimum content in distilled water is respectively：KNO385mg/L, MgSO4∙7H2O 150mg/L, KH2PO473mg/L,
Ca(NO3)2∙4H2O 180mg/L, MnSO4∙4H2O 27.5mg/L, Na2- EDTA 12.5mg/L, FeSO4∙7H2O 13.5mg/L,
H3BO38.25mg/L, ZnSO4∙7H2O 9.0mg/L, KI 0.5mg/L, AgNO317mg/L, Na2MoO4∙2H2O 0.25mg/L,
CuSO4∙5H2O 0.025mg/L, CoCl2∙6H2O 0.025mg/L, adenine 15mg/L, glutamine 500mg/L, vitamin
B1 1.0mg/L, vitamin B6 0.7mg/L, Vc 0.6mg/L, nicotinic acid 5.0mg/L, inositol 130mg/L, glycine
4.0mg/L, insulin 2.0mg/L, cyclic guanylic acid 0.2g/L, glutathione 32.5mg/L, ethylmethane sulfonate 1.2g/L,
Sucrose 45g/L, agar 7.0g/L, 6-BA 1.0mg/L, NAA 1.0mg/L, mannitol 15g/L, coconut palm breast 30g/L, plant sulphur
Change kinetin PSK- α 20pmol/L.
3. the muskmelon flower pesticide inducing culture according to claim 1 and 2, it is characterised in that：Described pyrovinic acid second
The adding method of ester is：Prepare after ethylmethane sulfonate solution, sterilized with 0.22 μm of filtering with microporous membrane of sterilizing, wait to train
When condensing to 45-55 DEG C after foster base high pressure steam sterilization, taking-up is placed in superclean bench, sterile working, adds pyrovinic acid
Ethyl ester solution, shakes up in condensation, 24h times and is inoculated with.
4. the muskmelon flower pesticide inducing culture according to claim 1 and 2, it is characterised in that：Described culture medium is matched somebody with somebody
PH value processed is：5.4-5.8, optimal secure ph is 5.6.
5. a kind of muskmelon Anther culture breeding method, it is characterised in that：Anther culture breeding method is operated as follows：
（1）Bud is drawn materials and pretreatment
When in crop field, the muskmelon of conventional cultivation is in initial bloom stage, in morning 9-11 points collection robust growth, the plant of no disease and pests harm
It is examination material to take developmental stage to be in the bud of mid-late uninucleate stage, and self-sealing plastic bag is used again after the bud of collection is wrapped up with wet gauze
Cover, be placed in low-temperature treatment 2d in 4 DEG C of refrigerators, be then transferred to Heat thermostability 2d under 36 DEG C of environment；
（2）The preparation of culture medium
It is sub-packed in after culture medium described in claim 1,2,3 and 4 is prepared in 250ml triangular flask, every bottle of Sheng 50ml-
Be placed in after 70ml culture medium, sealing temperature be 121 DEG C, pressure be the 20min that sterilizes under the conditions of 15kPa high steams, condense to
At 45-55 DEG C, taking-up is placed in superclean bench, sterile working, is added ethylmethane sulfonate solution, is shaken up condensation, 24h times
Interior inoculation；
（3）Flower pesticide is inoculated with
Pretreated bud is taken out, first 1-2 min is rinsed with flowing water, peels off anthocaulus with tweezers, use fritter gauze wrapped, is immersed
15s in 70% alcohol, is sterilized 8-10 minutes with being transferred to after aseptic water washing 2-3 times in 0.1% mercuric chloride solution, is then rushed with sterilized water
Wash 3-4 times, bud is put into the sterilizing culture dish for being covered with filter paper, flower pesticide is stripped under aseptic condition be inoculated into step 2 and prepare
Inducing culture on, every bottle access 14-16 pieces of flower pesticide；
（4）Flower pesticide Fiber differentiation
By be inoculated with flower pesticide after culture medium be placed in temperature for 26-28 DEG C, intensity of illumination 1400-1800lx, photoperiod 11h light/13h
Cultivated under dark environment, induce antherderived callus；
（5）Flower pesticide differentiation culture
After after antherderived callus length to diameter 2.5cm, select translucent, loosely organized callus and transfer on differential medium
Differentiation culture is carried out, callus gradually differentiates green seedling；
（6）Domestication is with transplanting
Green seedling growth is vigorous after flower pesticide differentiation culture 35d, takes out test tube seedling after sealed membrane now is opened into domestication hardening, 7d,
Root is cleaned up, greenhouse, conventional cultivation is transplanted into.
6. muskmelon Anther culture breeding method according to claim 5, it is characterised in that：The step（1）Mid-late uninucleate stage
Muskmelon bud selection standard be：The vertical stem length degree of muskmelon bud is measured, the bud that stem length degree is 3.0-3.6mm is indulged.
7. muskmelon Anther culture breeding method according to claim 5, it is characterised in that：The step（5）Break up the bar of culture
Part is：25-27 DEG C of temperature, humidity are 70%-80%, intensity of illumination 2000-2500lux, photoperiod 12h light/12h dark.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710589435.8A CN107155897B (en) | 2017-07-19 | 2017-07-19 | A kind of muskmelon anther induced medium and Anther culture breeding method |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710589435.8A CN107155897B (en) | 2017-07-19 | 2017-07-19 | A kind of muskmelon anther induced medium and Anther culture breeding method |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN107155897A true CN107155897A (en) | 2017-09-15 |
| CN107155897B CN107155897B (en) | 2019-02-22 |
Family
ID=59817994
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201710589435.8A Expired - Fee Related CN107155897B (en) | 2017-07-19 | 2017-07-19 | A kind of muskmelon anther induced medium and Anther culture breeding method |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN107155897B (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114158481A (en) * | 2021-12-27 | 2022-03-11 | 浙江宜格企业管理集团有限公司 | Preparation method of magnolia callus culture |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104686371A (en) * | 2015-04-04 | 2015-06-10 | 王开 | Efficient induction medium formula of sorghum anther |
| CN105660405A (en) * | 2016-02-16 | 2016-06-15 | 江苏强农农业技术服务有限公司 | Formula of inducing medium for tomato anther culture breeding |
| CN106489728A (en) * | 2015-09-08 | 2017-03-15 | 无锡南理工科技发展有限公司 | A kind of Anthers of Hordeum Vulgare inducing culture based formulas |
-
2017
- 2017-07-19 CN CN201710589435.8A patent/CN107155897B/en not_active Expired - Fee Related
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104686371A (en) * | 2015-04-04 | 2015-06-10 | 王开 | Efficient induction medium formula of sorghum anther |
| CN106489728A (en) * | 2015-09-08 | 2017-03-15 | 无锡南理工科技发展有限公司 | A kind of Anthers of Hordeum Vulgare inducing culture based formulas |
| CN105660405A (en) * | 2016-02-16 | 2016-06-15 | 江苏强农农业技术服务有限公司 | Formula of inducing medium for tomato anther culture breeding |
Non-Patent Citations (2)
| Title |
|---|
| 张传海等: "《植物生物技术》", 30 September 2015 * |
| 马刘峰: "甜瓜花药离体培养研究", 《中国优秀博硕士学位论文全文数据库（硕士）农业科技辑》 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114158481A (en) * | 2021-12-27 | 2022-03-11 | 浙江宜格企业管理集团有限公司 | Preparation method of magnolia callus culture |
Also Published As
| Publication number | Publication date |
|---|---|
| CN107155897B (en) | 2019-02-22 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN106171958B (en) | A kind of wheat scab disease resistant and breeding method | |
| CN105532480B (en) | A kind of potato flower pesticide inducing culture and Anther culture breeding method | |
| CN105941152B (en) | A kind of asparagus all-male breeding method | |
| CN103081807B (en) | Method for regenerating plant by use of callus of camellia japonica | |
| CN103168685B (en) | Method for cultivating rhizoma bletillae polyploidy plants | |
| CN106171957B (en) | A kind of wheat salt tolerance breeding method that excellent salt-resistance strain is screened using flower training monoploid | |
| CN109258468A (en) | A kind of potato saline-alkali tolerant improvement breeding method | |
| CN109197573A (en) | A kind of rapid breeding method of high yield and high quality oil tea | |
| CN111657146A (en) | Induction culture medium and induction culture method for culture breeding of green Chinese onion flowers | |
| CN103688860B (en) | Culture medium for rapid propagation and seedling of dendrobium officinale protocorm like-bodies and tissue culture method | |
| CN106508661A (en) | Method for constructing anoectochilus formosanus polyploidy resource | |
| CN103814823B (en) | Culture method for inducing angelica dahurica anther to seedlings | |
| CN108782252A (en) | A kind of method of micrografting in fresh water Chinese pear tissue-cultured seedling test tube | |
| CN103636506B (en) | method for performing plant culture by utilizing shepherdia argentea caulicle regenerated plant induction culture medium and | |
| CN107155897B (en) | A kind of muskmelon anther induced medium and Anther culture breeding method | |
| CN105660411B (en) | The tissue-culturing rapid propagation method for culturing seedlings of almond ringdove chrysanthemum | |
| CN108085363A (en) | One identification method to grow tobacco to fusarium tabacinum Resistance To Root Rot Disease | |
| CN112088775A (en) | Method for raising seedlings of hispid fig in test tube | |
| CN105850745B (en) | A kind of matrimony vine flower pesticide inducing culture and Anther culture breeding method | |
| CN109220781A (en) | A kind of Cucumber Target Leaf Spot resistance breeding method | |
| CN103125391B (en) | Local multi-generation breeding method for soybean | |
| KR101887221B1 (en) | Method of mass propagation of bamboo by in vitro culture | |
| CN109220782A (en) | A kind of cucumber fusarium axysporum resistance breeding method | |
| CN102119662B (en) | Polyploidy induction method for fiber linum usitatissimum under isolated culture condition | |
| CN112042543A (en) | Method for obtaining haploid plant through in vitro culture of pear unfertilized ovule |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| TA01 | Transfer of patent application right |
Effective date of registration: 20190109 Address after: 745600 No. 166 Huawu Road, Huachi County, Qingyang City, Gansu Province Applicant after: Huachi Information Industry Service Center Address before: 222000 Hengli house, Qingnian Road complex, Haizhou, Lianyungang, Jiangsu, 2 floor. Applicant before: JIANGSU QIANGNONG AGRICULTURAL TECHNOLOGY SERVICE CO.,LTD. |
|
| TA01 | Transfer of patent application right | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20190222 |
|
| CF01 | Termination of patent right due to non-payment of annual fee |