CN105638479A - Method for rapidly propagating large cherries - Google Patents
Method for rapidly propagating large cherries Download PDFInfo
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- CN105638479A CN105638479A CN201610076532.2A CN201610076532A CN105638479A CN 105638479 A CN105638479 A CN 105638479A CN 201610076532 A CN201610076532 A CN 201610076532A CN 105638479 A CN105638479 A CN 105638479A
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- alabastrum
- large cherry
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- 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
- A01H4/008—Methods for regeneration to complete plants
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Abstract
The invention relates to a method for rapidly propagating large cherries. The method includes the steps of selecting early-stage mononuclear anther for disinfecting; inoculating a callus inducing medium, and conducting culturing to obtain calluses; inoculating a differential medium, and conducting culturing to obtain test tube plantlets; inoculating a rooting medium, and conducting screening to obtain monomer plants. The inducing rate of the calluses of cultured large cherry pollen can be 88%, the rooting rate can be 96%, and the transplanting survival rate can be 98%.
Description
Technical field
The present invention relates to a kind of method for plant tissue culture, be specifically related to a kind of large cherry quick-breeding method.
Background technology
Fructus Pruni pseudocerasi belongs to Rosaceae deciduous tree fruit tree, in beingState northFruit maturation fruit tree species the earliest after side's deciduous fruit tree relaying cherry, therefore, early has the good reputation of " spring fruit first ". Regulating fresh fruit dull season, balanced year-round supply and meet the needs aspect of people's lives, there is special effect. Large cherry fruit contains more rich nutrient substance, and the health of people is had certain nutritive value. Chinese medicine and pharmacy is thought, big cherry is chosen has QI invigorating in tune, the function of wind-damp dispelling. Large cherry management recruitment is few, and production cost is low, high financial profit, be suitable in Liaoning, Shandong, Shaanxi, Henan, the cultivation of the ground such as Hebei.
Development along with plant tissue culture technique, plant cell has " totipotency " and is widely confirmed, therefore, flower pesticide is cultivated by vitro method in vitro when, the artificial development pathway changing sporidiole so that it is Development of Gametophytes approach terminates, and turns to sporinite to grow, occurred by organ or embryo occur approach, form complete plant. The report cultivated about large cherry in-vitro pollen is little, and inductivity, survival rate are low.
Summary of the invention
The present invention is directed to current problem, it is provided that a kind of large cherry quick-breeding method.
For solving above-mentioned technical problem, the technical scheme that the present invention adopts is: provide a kind of large cherry quick-breeding method, including,
1) alabastrum collection preserves: takes to grow of the right age alabastrum one strain of early stage or many strains without disease pest, well-grown monokaryon on large cherry plant inflorescence in fine day, is preserved with ice chest rapidly by the alabastrum taken;
2) flower pesticide sterilization: the alabastrum being saved in step 1 in ice chest is immersed on superclean bench sterilization 10-15s in the ethanol of 70-75% concentration, quickly remove and immerse sterilization more than 8min in saturated liquor natrii hypochloritis immediately, rocking beaker in disinfecting process gently makes it sterilize fully, is finally repeatedly placed on aseptic water washing on filter paper afterwards and blots alabastrum surface moisture;
3) flower pesticide in alabastrum is stripped, the flower pesticide inoculation callus inducing medium that then will strip, it is placed on shaking table and cultivates, make its pollen in the process of shake naturally become scattered about in culture medium, cultivate 10-20 days, choose anther wall, continue to cultivate, it is thus achieved that callus;
4) inoculation division culture medium, cultivates and obtains test tube seedling;
5) inoculation root media, screening obtains monomer plant.
Preferably, described callus inducing medium: 1/2MS+6-BA0.8mg/L+ZT0.6mg/L+NAA0.4��0.6mg/L+ glycine 1.1mg/L+ thiamine hydrochloride 1.2mg/L+ nicotinic acid 0.7mg/L+ agar 1000��2000mg/L+ sucrose 20��30g/L+ yeast extract 2000��3000mg/L, 18-20 DEG C, intensity of illumination be 1800LX, light application time 8 hour/day when cultivate.
Preferably, described division culture medium is 3/4MS+6-BA2.0mg/L+NAA0.5��0.7mg/L+7% sucrose, 20-22 DEG C, intensity of illumination be 2000LX, light application time 12 hour/day when cultivate.
Preferably, described root media be 1/2MS+IBA0.1��0.3mg/L+NAA0.3��0.5mg/L+ agar 40��60g/L+ sucrose 2000��3000mg/L+ VB11��3mg/L+ quality of activated carbon than 0.1%��0.3%+ yeast extract 5000��7000mg/L, 23-25 DEG C, intensity of illumination cultivates when be 2200LX, light application time being 15 hour/day.
Preferably, also include acclimatization and transplants, it is specially corkage seedling exercising 4��6d when long more than 5 roots, Seedling in bottle is taken out, shoot root base portion 5��8min is soaked with 0.003%��0.005% mass ratio IAA solution, tissue cultured seedling is transplanted in substrate, water foot and determine root water, covering Small plastic shed film heat and moisture preserving, early stage temperature is maintained at 15��20 DEG C, humidity 80��90%, film is taken off after 7��15d, progressively extend film-uncovering time, until not overlay film, watered every 2��3 weeks and execute 0.1%��0.3% weight ratio carbamide and 0.1%��0.3% mass ratio KH2PO4Or thin liquid dung, transplantation of seedlings will be survived; Described substrate is: leaf mould: perlite: Vermiculitum=2: 2: 1 volume ratios.
Preferably, during taking off film, mean temperature keeps 15 DEG C��25 DEG C, and humidity keeps 80%��95%.
The invention has the beneficial effects as follows: the present invention adopts large cherry flower pesticide to be outer implant, adjust the period of medicine of deflorating, carry out the induction of anther callus, adopt the method alternate culture that illumination cultivation and light culture combine, it is thus achieved that pollen plant. Culture medium callus proliferation of the present invention is obvious, is easily formed pollen plant. It is crucial that have found suitable cultural method, by the method for the present invention, the in vitro pollen of large cherry is cultivated, there is pollen germination and become that the embryo time is short, doubling etticiency is high, yield is big and is easier to obtain the advantages such as excellent variation breeding intermediate materials, at aspects such as breeding material purification and rejuvenation, Germplasm enhancement, polyploid breeding and pollen development process researchs, there is bigger practical value.
Detailed description of the invention
Below presently preferred embodiments of the present invention is described in detail, so that advantages and features of the invention can be easier to be readily appreciated by one skilled in the art, thus protection scope of the present invention being made apparent clear and definite defining.
The embodiment of the present invention includes:
Embodiment one: a kind of " early big fruit " large cherry quick-breeding method, including,
1) alabastrum collection preserves: takes to grow of the right age alabastrum one strain of early stage or many strains without disease pest, well-grown monokaryon on large cherry plant inflorescence in fine day, is preserved with ice chest rapidly by the alabastrum taken;
2) flower pesticide sterilization: the alabastrum being saved in step 1 in ice chest is immersed on superclean bench sterilization 15s in the ethanol of 75% concentration, quickly remove and immerse sterilization 10min in saturated liquor natrii hypochloritis immediately, rocking beaker in disinfecting process gently makes it sterilize fully, is finally repeatedly placed on aseptic water washing on filter paper afterwards and blots alabastrum surface moisture;
3) flower pesticide in alabastrum is stripped, the flower pesticide inoculation callus inducing medium that then will strip, it is placed on shaking table and cultivates, make its pollen in the process of shake naturally become scattered about in culture medium, cultivate 15 days, choose anther wall, continue to cultivate, it is thus achieved that callus; Described callus inducing medium: 1/2MS+6-BA0.8mg/L+ZT0.6mg/L+NAA0.5mg/L+ glycine 1.1mg/L+ thiamine hydrochloride 1.2mg/L+ nicotinic acid 0.7mg/L+ agar 1500mg/L+ sucrose 25g/L+ yeast extract 2500mg/L, 18 DEG C, intensity of illumination be 1800LX, light application time 8 hour/day when cultivate.
4) inoculation division culture medium, cultivates and obtains test tube seedling; Described division culture medium is 3/4MS+6-BA2.0mg/L+NAA0.6mg/L+7% sucrose, 22 DEG C, intensity of illumination be 2000LX, light application time 12 hour/day when cultivate.
5) inoculation root media, screening obtains monomer plant, described root media be 1/2MS+IBA0.2mg/L+NAA0.4mg/L+ agar 50g/L+ sucrose 2500mg/L+ vitamin B12 mg/L+ quality of activated carbon than 0.2%+ yeast extract 6000mg/L, 25 DEG C, intensity of illumination cultivates when be 2200LX, light application time being 15 hour/day.
Acclimatization and transplants, is specially corkage seedling exercising 5d when long more than 5 roots, is taken out by Seedling in bottle, shoot root base portion 6min is soaked with 0.004% mass ratio IAA solution, tissue cultured seedling is transplanted in substrate, waters foot and determine root water, cover Small plastic shed film heat and moisture preserving, early stage temperature is maintained at 18 DEG C, humidity 85%, takes off film after 10d, progressively extend film-uncovering time, until not overlay film, watered every 2 weeks and execute 0.2% mass than carbamide and 0.2% mass ratio KH2PO4Or thin liquid dung, transplantation of seedlings will be survived; Described substrate is: leaf mould: perlite: Vermiculitum=2: 2: 1 volume ratios. During taking off film, mean temperature keeps 20 DEG C, and humidity keeps 80%��95%.
Result through above-mentioned cultivation is that the inductivity of callus reaches 85%, and rooting rate reaches 95%, and transplanting survival rate reaches 99%.
Embodiment two: a kind of " early big fruit " large cherry quick-breeding method, including,
1) alabastrum collection preserves: takes to grow of the right age alabastrum one strain of early stage or many strains without disease pest, well-grown monokaryon on large cherry plant inflorescence in fine day, is preserved with ice chest rapidly by the alabastrum taken;
2) flower pesticide sterilization: the alabastrum being saved in step 1 in ice chest is immersed on superclean bench sterilization 15s in the ethanol of 75% concentration, quickly remove and immerse sterilization more than 8min in saturated liquor natrii hypochloritis immediately, rocking beaker in disinfecting process gently makes it sterilize fully, is finally repeatedly placed on aseptic water washing on filter paper afterwards and blots alabastrum surface moisture;
3) flower pesticide in alabastrum is stripped, the flower pesticide inoculation callus inducing medium that then will strip, it is placed on shaking table and cultivates, make its pollen in the process of shake naturally become scattered about in culture medium, cultivate 20 days, choose anther wall, continue to cultivate, it is thus achieved that callus; Described callus inducing medium: 1/2MS+6-BA0.8mg/L+ZT0.6mg/L+NAA0.6mg/L+ glycine 1.1mg/L+ thiamine hydrochloride 1.2mg/L+ nicotinic acid 0.7mg/L+ agar 2000mg/L+ sucrose 30g/L+ yeast extract 3000mg/L, 20 DEG C, intensity of illumination be 1800LX, light application time 8 hour/day when cultivate.
4) inoculation division culture medium, cultivate obtain test tube seedling, described division culture medium is 3/4MS+6-BA2.0mg/L+NAA0.7mg/L+7% sucrose, 21 DEG C, intensity of illumination be 2000LX, light application time 12 hour/day when cultivate;
5) inoculation root media, screening obtains monomer plant, described root media be 1/2MS+IBA0.3mg/L+NAA0.5mg/L+ agar 60g/L+ sucrose 3000mg/L+ orotic acid mg/L+ quality of activated carbon than 0.3%+ yeast extract 7000mg/L, 25 DEG C, intensity of illumination cultivates when be 2200LX, light application time being 15 hour/day.
Acclimatization and transplants, is specially corkage seedling exercising 6d when long more than 5 roots, is taken out by Seedling in bottle, shoot root base portion 8min is soaked with 0.005% weight ratio IAA solution, tissue cultured seedling is transplanted in substrate, waters foot and determine root water, cover Small plastic shed film heat and moisture preserving, early stage temperature is maintained at 15 DEG C, humidity 90%, takes off film after 15d, progressively extend film-uncovering time, until not overlay film, watered every 3 weeks and execute 0.3% weight ratio carbamide and 0.3% weight ratio KH2PO4Or thin liquid dung, transplantation of seedlings will be survived;Described substrate is: leaf mould: perlite: Vermiculitum=2: 2: 1 volume ratios. During taking off film, mean temperature keeps 25 DEG C, and humidity keeps 95%.
Result through above-mentioned cultivation is that the inductivity of callus reaches 88%, and rooting rate reaches 96%, and transplanting survival rate reaches 98%.
The foregoing is only embodiments of the invention; not thereby the scope of the claims of the present invention is limited; every equivalent structure utilizing description of the present invention to make or equivalence flow process conversion; or directly or indirectly it is used in other relevant technical fields, all in like manner include in the scope of patent protection of the present invention.
Claims (6)
1. a large cherry quick-breeding method, it is characterised in that comprise the following steps:
1) alabastrum collection preserves: takes to grow of the right age alabastrum one strain of early stage or many strains without disease pest, well-grown monokaryon on large cherry plant inflorescence in fine day, is preserved with ice chest rapidly by the alabastrum taken;
2) flower pesticide sterilization: the alabastrum being saved in step 1 in ice chest is immersed on superclean bench sterilization 10-15s in the ethanol of 70-75% concentration, quickly remove and immerse sterilization more than 8min in saturated liquor natrii hypochloritis immediately, rocking beaker in disinfecting process gently makes it sterilize fully, is finally repeatedly placed on aseptic water washing on filter paper afterwards and blots alabastrum surface moisture;
3) flower pesticide in alabastrum is stripped, the flower pesticide inoculation callus inducing medium that then will strip, it is placed on shaking table and cultivates, make its pollen in the process of shake naturally become scattered about in culture medium, cultivate 10-20 days, choose anther wall, continue to cultivate, it is thus achieved that callus;
4) inoculation division culture medium, cultivates and obtains test tube seedling;
5) inoculation root media, screening obtains monomer plant.
2. large cherry quick-breeding method according to claim 1, it is characterized in that, described callus inducing medium: 1/2MS+6-BA0.8mg/L+ZT0.6mg/L+NAA0.4��0.6mg/L+ glycine 1.1mg/L+ thiamine hydrochloride 1.2mg/L+ nicotinic acid 0.7mg/L+ agar 1000��2000mg/L+ sucrose 20��30g/L+ yeast extract 2000��3000mg/L, 18-20 DEG C, intensity of illumination be 1800LX, light application time 8 hour/day when cultivate.
3. the large cherry quick-breeding method as described in any one of claim 1-2, it is characterized in that, described division culture medium is 3/4MS+6-BA2.0mg/L+NAA0.5��0.7mg/L+7% sucrose, 20-22 DEG C, intensity of illumination be 2000LX, light application time 12 hour/day when cultivate.
4. the large cherry quick-breeding method as described in any one of claim 1-3, it is characterized in that, described root media be 1/2MS+IBA0.1��0.3mg/L+NAA0.3��0.5mg/L+ agar 40��60g/L+ sucrose 2000��3000mg/L+ VB11��3mg/L+ quality of activated carbon than 0.1%��0.3%+ yeast extract 5000��7000mg/L, 23-25 DEG C, intensity of illumination cultivates when be 2200LX, light application time being 15 hour/day.
5. the large cherry quick-breeding method as described in any one of claim 1-4, it is characterized in that, also include acclimatization and transplants, it is specially corkage seedling exercising 4��6d when long more than 5 roots, Seedling in bottle is taken out, shoot root base portion 5��8min is soaked with 0.003%��0.005% mass ratio IAA solution, tissue cultured seedling is transplanted in substrate, water foot and determine root water, cover Small plastic shed film heat and moisture preserving, early stage temperature is maintained at 15��20 DEG C, humidity 80��90%, film is taken off after 7��15d, progressively extend film-uncovering time, until not overlay film, watered every 2��3 weeks and execute 0.1%��0.3% weight ratio carbamide and 0.1%��0.3% weight ratio KH2PO4Or thin liquid dung, transplantation of seedlings will be survived;Described substrate is: leaf mould: perlite: Vermiculitum=2: 2: 1 volume ratios.
6. large cherry quick-breeding method according to claim 5, it is characterised in that during taking off film, mean temperature keeps 15 DEG C��25 DEG C, and humidity keeps 80%��95%.
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| CN201610076532.2A CN105638479A (en) | 2016-02-03 | 2016-02-03 | Method for rapidly propagating large cherries |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107494543A (en) * | 2017-08-01 | 2017-12-22 | 山东大丰园农业有限公司 | A kind of root-growing agent and its outside sprout-cultivating-bottle method for cherry tissue culture outside sprout-cultivating-bottle radication |
| CN108834908A (en) * | 2018-09-10 | 2018-11-20 | 河北农业大学 | Peach anther callus induction and subculture preservation method |
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| KR20150026554A (en) * | 2013-09-03 | 2015-03-11 | 대한민국(관리부서 : 산림청 국립산림과학원장) | Development of a micropropagation technique in Prunus avium clones |
| CN104737919A (en) * | 2015-04-24 | 2015-07-01 | 芜湖东源新农村开发股份有限公司 | Tissue culture rapid seedlings growing method used for cherry stock M9 |
| CN105154384A (en) * | 2015-09-18 | 2015-12-16 | 江苏丘陵地区镇江农业科学研究所 | Method for improving large cherry pollen germination rate |
-
2016
- 2016-02-03 CN CN201610076532.2A patent/CN105638479A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR20150026554A (en) * | 2013-09-03 | 2015-03-11 | 대한민국(관리부서 : 산림청 국립산림과학원장) | Development of a micropropagation technique in Prunus avium clones |
| CN104737919A (en) * | 2015-04-24 | 2015-07-01 | 芜湖东源新农村开发股份有限公司 | Tissue culture rapid seedlings growing method used for cherry stock M9 |
| CN105154384A (en) * | 2015-09-18 | 2015-12-16 | 江苏丘陵地区镇江农业科学研究所 | Method for improving large cherry pollen germination rate |
Non-Patent Citations (2)
| Title |
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| CHRISTOPHER M. LONG ET.AL.,: "Production of a Microspore-derived Callus Population from Sweet Cherry", 《HORTSCIENCE》 * |
| 李晓燕等: "甜樱桃巨红13-38 花药发育时期鉴定及愈伤诱导的初步研究", 《大连民族学院学报》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107494543A (en) * | 2017-08-01 | 2017-12-22 | 山东大丰园农业有限公司 | A kind of root-growing agent and its outside sprout-cultivating-bottle method for cherry tissue culture outside sprout-cultivating-bottle radication |
| CN108834908A (en) * | 2018-09-10 | 2018-11-20 | 河北农业大学 | Peach anther callus induction and subculture preservation method |
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