CN104273034A - Induction and differentiation culture technique of Lycium ruthenicum Murr. - Google Patents
Induction and differentiation culture technique of Lycium ruthenicum Murr. Download PDFInfo
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- CN104273034A CN104273034A CN201410568523.6A CN201410568523A CN104273034A CN 104273034 A CN104273034 A CN 104273034A CN 201410568523 A CN201410568523 A CN 201410568523A CN 104273034 A CN104273034 A CN 104273034A
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Abstract
The invention discloses an induction and differentiation culture technique of Lycium ruthenicum Murr. and belongs to the technical field of plant breeding. The method not only can rapidly breed a large amount of high-quality seedlings for production practice but also can provide a technical support for improving the cultivation of new variety of Lycium ruthenicum Murr. by a biotechnology, and also is an idler approach to conservation of germplasm resources of Lycium ruthenicum Murr. The method comprises the following steps: selecting an annual robust and disease-free branch, shearing to be a stem with 1-2 axillary buds, disinfecting and inoculating onto an inductive bud growth medium, then transferring to a callus induction culture medium after new buds grow out, inducing to produce callus, transferring the callus to the induced differentiation medium to induce growth of cluster buds, carrying out rooting culture on the cluster buds; and finally acclimatizing the cultured rooted seedlings and transplanting.
Description
Technical field
The present invention relates to technical field of plant propagation, particularly relate to a kind of induction and differentiation culture technique of black fruit lyceum callus.
Background technology
Black fruit lyceum (Lycium ruthenicum Murr.) is Solanaceae (Solanceae) Lycium (Lycium L.) perennial shrub, be distributed in Xinjiang, Tibet, Qinghai, Gansu, Ningxia, North Shaanxi, also there is distribution in the Central Asia, Caucasia and Europe.Often be born in various salinization soil or the desert environments such as saline-alkali wasteland, salinization sand ground, bank, salt lake, roadside with shrubbery shape, there is very high photosynthetic efficiency and stronger resistance, the distinctive anti-salt drought resisting wild plant of NORTHWEST CHINA, even if extensive cultivation, also very easily artificial planting.Black fruit lyceum is except having the chemical composition similar to lycium barbarum (Lyciumbarbarum L.), and its fruit pigment is relatively stable, can be widely used in light industry, weaving, medicine and other fields.Black fruit lyceum has the plurality of advantages such as drought resisting, cold-resistant, strong, the resistance to soil depletion of Salt And Alkali Tolerance, root turion and nutritive value are high, be Saline-alkali Field Control, bank protection, the good seeds that prevent erosion, there is economic outlook and potentiality to be exploited widely.At saline-alkali wasteland and arid area severe water and soil erosion Regional cultivation, can local environment be improved, also can provide certain income for the local masses.Black fruit lyceum is in wild state at present substantially, and Proterozoic natural conditions are severe, and vegetation degeneration is serious, dead even in flakes in some area, causes very large threat to the growth of black fruit lyceum, protection and this vegetation resources of Appropriate application extremely urgent.
Callus tissue culture serves many purposes.Mechanism, the genetic variation rule of growth and development of plants and differentiation can be studied on the one hand, plant genetics and breeding is acquired a special sense.New variety of plant and stock breeding speed can be accelerated, rapid high efficiency, low cost; Cultivate virus-free nursery stock; Rapid high efficiency, low cost obtains the different plant of ploidy, overcomes distant hybridization difficulty; Be beneficial to germ plasm resource to preserve for a long time and long distance transportation; Manufacture of intraocular seed.Can be used for the useful compound of large-scale industrialized production on the other hand, or produce upper useful clone for cell chulture screening industry, agricultural, medicine, or for the protoplast source etc. in Protoplast cuhnre.Facts have proved, callus tissue culture is not only a kind of new tool of plant fast propagation, is also the desirable route of plant improvement, preserving seed and useful production of chemicals simultaneously.
Summary of the invention
The object of this invention is to provide a kind of induction and differentiation culture technique of black fruit lyceum callus.
For achieving the above object, the following technical scheme of employing of the present invention:
An induction and differentiation culture technique for black fruit lyceum callus, comprises the following steps: healthy and strong at experimental field growth selection, yield positive results fine individual plant annotinous branch many; With flowing water, branch is rinsed well, the branch rinsed is cut into 1-2 axillary bud stem section, and is placed on superclean bench, with the alcohol-pickled 2-5s of 75%, then use the mercuric chloride process 10min of 0.1%; Be inoculated on superclean bench on inducing culture by the material handled well, axillalry bud starts to sprout, and grows sprouting, and inducing culture is: MS+IBA 0.2mg/L; With new tooth blade for material, on superclean bench, blade is cut into the fritter of 0.5cm-1.0cm, received induction on callus inducing medium and produce callus, callus inducing medium is: MS+6-BA 1mg/L+NAA 1mg/L; Described callus is inoculated into induction on callus induction differential medium and produces Multiple Buds, callus induction differential medium is: MS+6-BA 0.2mg/L; By described Multiple Buds on superclean bench, become with 1-2 axillary bud stem section by scissor cut, be inoculated on root media, root media is: MS+6-BA 0.4mg/L+NAA 0.2mg/L; By the test-tube plantlet of root induction, in laboratory, open bottleneck hardening 7d, transplant in greenhouse; By in nutritive cube porose bottom acclimatization and transplants to the black polyethylene film of 8cm × 12cm.
Matrix in described nutritive cube is the ratio of forest soil, river sand and vermiculite is 2:1:1.Matrix be exposed to the sun under the sun after with 0.1% potassium permanganate sterilization.Lightly test-tube plantlet is taken out from triangular flask with tweezers, clean up the medium that root is residual.Be transplanted in ready container.Root is put in the duck eye accomplished fluently in advance during transplanting, root system is unfolded, compacting, make root system and soil close contact, prevent from planting dark, nest root and leak root, waters permeable after transplanting.For preventing damage by disease and insect, transplanting sprayed the carbendazim of 800-1000 many times the same day once, later according to circumstances at the suitable agricultural chemicals of sprinkling.After this, sprinkling irrigation is adopted to water according to actual conditions.When height of seedling 10-20cm, land for growing field crops can be transplanted to.
Owing to also not cultivating excellent black fruit lyceum new varieties at present, described experiment material is that the black fruit lyceum of the different regions, Gansu of collecting is planted in experimental field, through the fine individual plant selected by the observations of 2 years.Described medium all adds white sugar 20g/L, agar powder 4.5g/L, pH value 5.8-6.0 for reducing costs.The condition of culture of culturing room is: temperature about 28 DEG C (using airconditioning control temperature), and illumination every day 14-16h, intensity of illumination is 1500-3000lx.
The invention has the advantages that:
1. the induction and differentiation culture technique of a kind of black fruit lyceum callus provided by the invention can be a large amount of high quality seedling of production practices Fast-propagation;
2. bio-technology improvement black fruit lyceum also can be utilized further to cultivate new varieties technical support is provided;
3. be also a kind of desirable route of black fruit lyceum Germ-plasma resources protection simultaneously.
Accompanying drawing explanation
Fig. 1 is the incubation schematic diagram of the induction and differentiation culture technique of a kind of black fruit lyceum callus provided by the invention.
Embodiment
Below in conjunction with the accompanying drawing in the embodiment of the present invention, technical scheme of the present invention is clearly and completely described.
The invention provides a kind of induction and differentiation culture technique of black fruit lyceum callus, described method comprises:
(1) explant is selected: healthy and strong at experimental field growth selection, yield positive results fine individual plant annotinous branch many.
(2) explant process: branch is rinsed well at use for laboratory flowing water, the branch rinsed cuts into 1-2 axillary bud stem section, the stem section cleaned up, on superclean bench, with the alcohol-pickled 2-5s of 75%, then uses the mercuric chloride process 10min of 0.1% for subsequent use.
(3) Initial culture: material sterile-processed for step (2) is inoculated on inducing culture on superclean bench, and axillalry bud starts to sprout, and grows sprouting, and inducing culture is: MS+IBA 0.2mg/L.
(4) evoked callus: with the blade of the test-tube plantlet of step (3) for material, on superclean bench, blade is cut into the fritter of 0.5cm-1.0cm, received induction on callus inducing medium and produce callus, callus inducing medium is: MS+6-BA 1mg/L+NAA 1mg/L.
(5) Calli Differentiation: the callus that step (4) produces is inoculated into induction on callus induction differential medium and produces Multiple Buds, callus induction differential medium is: MS+6-BA 0.2mg/L.
(6) culture of rootage: by step (5) Multiple Buds on superclean bench, becomes with 1-2 axillary bud stem section by scissor cut, is inoculated on root media.Root media is: MS+6-BA 0.4mg/L+NAA 0.2mg/L.
(7) test-tube seedling transplanting: by the test-tube plantlet of step (6) root induction, open bottleneck hardening 7d in laboratory; Transplant in greenhouse.
(8) container nursery: in the nutritive cube that just step (7) transplantation of seedlings of refining is porose bottom the black polyethylene film of 8cm × 12cm, matrix in nutritive cube is forest soil, the ratio of river sand and vermiculite is 2:1:1, matrix be exposed to the sun under the sun after with 0.1% potassium permanganate sterilization, lightly test-tube plantlet is taken out from triangular flask with tweezers, clean up the medium that root is residual, be transplanted in ready container, during transplanting, root is put in the duck eye accomplished fluently in advance, root system is unfolded, compacting, make root system and soil close contact, prevent from planting dark, nest root and leakage root, water permeable after transplanting, for preventing damage by disease and insect, transplanting sprayed the carbendazim of 800-1000 many times the same day once, later according to circumstances at the suitable agricultural chemicals of sprinkling, after this, sprinkling irrigation is adopted to water according to actual conditions, when height of seedling 10-20cm, land for growing field crops can be transplanted to.
Claims (4)
1. an induction and differentiation culture technique for black fruit lyceum callus, is characterized in that: comprise the following steps:
(1) explant is selected: healthy and strong at experimental field growth selection, yield positive results fine individual plant annotinous branch many.
(2) explant process: branch is rinsed well at use for laboratory flowing water.The branch rinsed cuts into 1-2 axillary bud stem section, and the stem section cleaned up, on superclean bench, with the alcohol-pickled 2-5s of 75%, then uses the mercuric chloride process 10min of 0.1% for subsequent use.
(3) Initial culture: material sterile-processed for step (2) is inoculated on inducing culture on superclean bench, and axillalry bud starts to sprout, and grows sprouting, and inducing culture is: MS+IBA0.2mg/L.
(4) evoked callus: with the blade of the test-tube plantlet of step (3) for material, on superclean bench, blade is cut into the fritter of 0.5cm-1.0cm, received induction on callus inducing medium and produce callus, callus inducing medium is: MS+6-BA1mg/L+NAA1mg/L.
(5) Calli Differentiation: the callus that step (4) produces is inoculated into induction on callus induction differential medium and produces Multiple Buds, callus induction differential medium is: MS+6-BA0.2mg/L.
(6) culture of rootage: by step (5) Multiple Buds on superclean bench, becomes with 1-2 axillary bud stem section by scissor cut, is inoculated on root media.Root media is: MS+6-BA0.4mg/L+NAA0.2mg/L.
(7) test-tube seedling transplanting: by the test-tube plantlet of step (6) root induction, open bottleneck hardening 7d in laboratory; Transplant in greenhouse.
(8) container nursery: in the nutritive cube that just step (7) transplantation of seedlings of refining is porose bottom the black polyethylene film of 8cm × 12cm.
2. the induction and differentiation culture technique of a kind of black fruit lyceum callus according to claim 1, is characterized in that: medium all adds white sugar 20g/L, agar powder 4.5g/L, pH value 5.8-6.0 for reducing costs.
3. the induction and differentiation culture technique of a kind of black fruit lyceum callus according to claim 1 and 2, it is characterized in that: the condition of culture of culturing room is: temperature 27-29 DEG C (using airconditioning control temperature), illumination every day 14-16h, intensity of illumination is 1500-3000lx.
4. according to claim 1, the induction and differentiation culture technique of a kind of black fruit lyceum callus described in 2 and 3, it is characterized in that: the matrix in nutritive cube is the ratio of forest soil, river sand and vermiculite is 2:1:1, matrix be exposed to the sun under the sun after with 0.1% potassium permanganate sterilization.
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Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104719165A (en) * | 2015-03-30 | 2015-06-24 | 四川禾木本业农林科技有限公司 | Rapid tissue culture method for lycium ruthenicum murr |
| CN105325147A (en) * | 2015-10-30 | 2016-02-17 | 山西火品生物科技有限公司 | Method for cultivating and planting lycium ruthenicum murray |
| CN105325148A (en) * | 2015-10-30 | 2016-02-17 | 山西火品生物科技有限公司 | Wild lycium ruthenicum artificial domesticating planting method |
| CN105684901A (en) * | 2016-01-22 | 2016-06-22 | 甘肃省治沙研究所 | Quick propagation method for medicinal plant lycium ruthenicum in deserts |
| CN107135950A (en) * | 2017-06-19 | 2017-09-08 | 南京晓庄学院 | A kind of breeding method of quick acquisition black fruit fructus lycii regrowth |
| CN108605786A (en) * | 2018-04-10 | 2018-10-02 | 宁夏农林科学院枸杞工程技术研究所 | A kind of method that old stake branch root of hair makes matrimony vine potted landscape |
| CN109287480A (en) * | 2018-10-19 | 2019-02-01 | 宁夏农林科学院枸杞工程技术研究所 | A kind of method that black fruit fructus lycii Anther Culture obtains haplobiont |
| CN112753578A (en) * | 2021-01-25 | 2021-05-07 | 中国林业科学研究院林业研究所 | Method for inducing and rapidly proliferating Chinese wolfberry callus |
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Cited By (11)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104719165A (en) * | 2015-03-30 | 2015-06-24 | 四川禾木本业农林科技有限公司 | Rapid tissue culture method for lycium ruthenicum murr |
| CN104719165B (en) * | 2015-03-30 | 2016-08-17 | 四川禾木本业农林科技有限公司 | A kind of quickly tissue culture method of black fruit lyceum |
| CN105325147A (en) * | 2015-10-30 | 2016-02-17 | 山西火品生物科技有限公司 | Method for cultivating and planting lycium ruthenicum murray |
| CN105325148A (en) * | 2015-10-30 | 2016-02-17 | 山西火品生物科技有限公司 | Wild lycium ruthenicum artificial domesticating planting method |
| CN105684901A (en) * | 2016-01-22 | 2016-06-22 | 甘肃省治沙研究所 | Quick propagation method for medicinal plant lycium ruthenicum in deserts |
| CN105684901B (en) * | 2016-01-22 | 2018-07-13 | 甘肃省治沙研究所 | A kind of rapid propagation method of Desert Regions medicinal plant black fruit fructus lycii |
| CN107135950A (en) * | 2017-06-19 | 2017-09-08 | 南京晓庄学院 | A kind of breeding method of quick acquisition black fruit fructus lycii regrowth |
| CN108605786A (en) * | 2018-04-10 | 2018-10-02 | 宁夏农林科学院枸杞工程技术研究所 | A kind of method that old stake branch root of hair makes matrimony vine potted landscape |
| CN108605786B (en) * | 2018-04-10 | 2019-11-08 | 宁夏农林科学院枸杞工程技术研究所 | A kind of method of old stake branch root of hair production fructus lycii potted landscape |
| CN109287480A (en) * | 2018-10-19 | 2019-02-01 | 宁夏农林科学院枸杞工程技术研究所 | A kind of method that black fruit fructus lycii Anther Culture obtains haplobiont |
| CN112753578A (en) * | 2021-01-25 | 2021-05-07 | 中国林业科学研究院林业研究所 | Method for inducing and rapidly proliferating Chinese wolfberry callus |
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