CN111837953A - Rapid breeding method of sealwort seedlings - Google Patents
Rapid breeding method of sealwort seedlings Download PDFInfo
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- CN111837953A CN111837953A CN202010710730.6A CN202010710730A CN111837953A CN 111837953 A CN111837953 A CN 111837953A CN 202010710730 A CN202010710730 A CN 202010710730A CN 111837953 A CN111837953 A CN 111837953A
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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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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G22/00—Cultivation of specific crops or plants not otherwise provided for
- A01G22/25—Root crops, e.g. potatoes, yams, beet or wasabi
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G9/00—Cultivation in receptacles, forcing-frames or greenhouses; Edging for beds, lawn or the like
- A01G9/02—Receptacles, e.g. flower-pots or boxes; Glasses for cultivating flowers
- A01G9/029—Receptacles for seedlings
- A01G9/0299—Handling or transporting of soil blocks or seedlings
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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
- A01H4/001—Culture apparatus for tissue culture
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Abstract
The invention provides a rapid breeding method of polygonatum sibiricum seedlings, which comprises the following steps: 1) obtaining sealwort tissue culture tubers; 2) carrying out tissue culture on the sealwort tissue culture tubers by using a tissue culture seedling breeding technology to obtain tissue culture seedlings; 3) and (3) carrying out hardening culture on the tissue culture seedlings by utilizing a tissue culture hardening-seedling technology until the diameter of the hardened-seedling rhizome is more than or equal to 2.5cm, and obtaining the required seedlings. The method of combining the tuber tissue culture breeding technology with the seedling hardening-off technology is adopted, the problems of long breeding time and low seedling standardization degree are solved, the problems of long seedling time, irregular seedling emergence, seed provenance mixing and the like in the direct seeding and seedling culture of the sealwort seeds are solved, and in addition, the pure positive of the provenance can be guaranteed because the selected material for tissue culture is a rhizome.
Description
Technical Field
The invention belongs to the technical field of plant culture, and particularly relates to a rapid breeding method of polygonatum sibiricum seedlings.
Background
Rhizoma Polygonati (Polygonatum Rhizoma) is a perennial herb of Polygonatum of Liliaceae, is also called as "Xianren remainder food", is a traditional Chinese medicinal material in China, and has the effects of invigorating qi, nourishing yin, invigorating spleen, moistening lung and tonifying kidney. The "chinese pharmacopoeia" (2015 edition) contains dried rhizome of polygonatum kingianum (Polygonatum kingianum Coll. et Hemsl.), polygonatum (Polygonatum bicucum Delar. ex Redoute) and Polygonatum cyrtonema Hua (Polygonatum cyrtonema Hua), which are known as "rheum emodin", "Polygonatum Candidum" and "Polygonatum Candidum rhizome", according to different properties. The sealwort is widely distributed, and species differences of the plants tend to be complex due to morphological transition and geographical distribution overlapping, the species division is difficult, and the species intermediate types are more. Rhizoma polygonati is a traditional Chinese medicinal material with homology of medicine and food, has excellent nourishing efficacy, is a good product for prolonging life and nourishing body and protecting health since ancient times, and particularly, the demand of rhizoma polygonati is greatly increased year by year along with the development of rhizoma polygonati food and health care products in recent years.
At present, the sealwort seedling breeding technology is mainly a breeding method for direct seeding and seedling raising of seeds. The sealwort seed is a recalcitrant seed, the germination of the seed has certain limitation, the seedling breeding technology of direct seeding of the seed is limited, although the breeding technology is continuously broken through at present, the basic requirement of seedling transplanting of direct seeding of the seed is that the diameter of the root and stem of the seedling is more than or equal to 2.5cm, the seedling raising time when the diameter of the root and stem of the seedling directly seeded by the seed reaches 2.5cm needs 2 years, and the seedling can emerge in the second year after about 30% of the seeds are sowed. Therefore, the method for breeding the sealwort by directly seeding the seeds as the seedlings has low efficiency and is not suitable for large-scale standardized production. The problems of long seedling raising time, low seed emergence rate and seed source mixing exist in the direct seeding of seeds, and the planting cost of polygonatum sibiricum is greatly increased. In order to solve the problems, the invention provides a rapid breeding method of polygonatum sibiricum seedlings.
Disclosure of Invention
Aiming at the problems, the invention provides a rapid breeding method of polygonatum sibiricum seedlings, which comprises the following steps:
1) obtaining sealwort tissue culture tubers;
2) carrying out tissue culture on the sealwort tissue culture tubers by using a tissue culture seedling breeding technology to obtain tissue culture seedlings;
3) And (3) carrying out hardening culture on the tissue culture seedlings by utilizing a tissue culture hardening-seedling technology until the diameter of the hardened-seedling rhizome is more than or equal to 2.5cm, and obtaining the required seedlings.
Further, the obtaining of the sealwort tissue culture tuber comprises the following steps:
obtaining rhizome of polygonatum sibiricum with a confirmed seed source, wherein the rhizome of polygonatum sibiricum is a fresh rhizome of polygonatum sibiricum which is strong in growth, free of rot, free of plant diseases and insect pests and provided with buds and buds, and removing dead branches and fibrous roots to obtain rhizome tubers;
cleaning the obtained rhizoma Polygonati tuber, washing with detergent for 5-8min, and washing with running water for 2-3 h; sterilizing the cleaned rhizoma Polygonati tuber with 75% alcohol for 25-35s, washing with sterile water for 2-3 times, sterilizing with 0.2% mercuric chloride for 10-15min, and cleaning with sterile water for 6-7 times;
cutting rhizoma Polygonati stem with bud into 1-3cm diameter rhizoma Polygonati tissue culture tuber.
Further, the tissue culture cultivation of the sealwort tissue culture tuber by using the tissue culture seedling breeding technology comprises the following steps:
inoculating the sealwort tissue culture tubers on a first culture medium, and culturing for 7-14 days in a culture room to obtain cluster buds;
removing branches and leaves of the cluster buds, cutting the cluster buds into tubers with the diameter of 1cm, inoculating the tubers on a second culture medium, and culturing in a culture room for 20-30 days to obtain subculture cluster buds;
removing branches and leaves of the subculture multiple buds, cutting into tubers with the diameter of 1cm, inoculating the tubers on a third culture medium, and culturing in a culture room for 30d to obtain proliferation multiple buds;
Removing branches and leaves of the proliferated multiple buds, cutting the multiple buds into tubers with the diameter of 1cm, and inoculating the tubers onto a fourth culture medium; culturing in a culture room for 30 days to obtain differentiated cluster buds;
and cutting the cluster buds into tubers with buds, inoculating the tubers with the buds onto a fifth culture medium, and culturing for 60 days in a culture chamber to obtain tissue culture seedlings with roots.
Further, the culture conditions of the culture chamber are: at 25 ℃, the light is 10h, the dark is 14h, and the light intensity is 6000 lux.
Still further, the first medium is: MS is taken as a basic culture medium, and a culture medium of 3.0mg/LBA, 1.0mg/LNAA, 30g/L sucrose and 5g/L agar is added.
Still further, the second medium is: MS is taken as a basic culture medium, and a culture medium of 4.0mg/L BA, 0.5mg/L2,4-D, 30g/L sucrose and 5g/L agar is added.
Still further, the third medium is: MS is taken as a basic culture medium, and a culture medium of 2.0mg/L BA, 0.5mg/L LNAA, 30g/L sucrose and 5g/L agar is added.
Still further, the fourth medium is: MS is taken as a basic culture medium, and a culture medium of 1.0mg/L BA, 0.2mg/L LNAA, 30g/L sucrose and 5g/L agar is added.
Still further, the fifth medium is: 1/2MS is used as a basic culture medium, and a culture medium of 1.0mg/L NAA, 0.5g/L active carbon, 30g/L sucrose and 5g/L agar is added.
Further, the tissue culture seedling exercising culture by using the tissue culture seedling exercising technology comprises the following steps:
taking out the rooted aseptic seedlings, dressing the seedlings, transplanting the seedlings into a mixed matrix, and turning green the seedlings after 30 days;
spraying the mixed solution A after the aseptic seedlings are transplanted to the matrix for 40 days; spraying the mixed solution B after 60 days, and hardening seedlings after 90 days to obtain sealwort seedlings.
Still further, the seed dressing comprises: putting the sterile seedlings of the clean culture medium into a container, and adding the mixture of the components in percentage by mass as carbendazim: iprodione: thiram is 1: 1: 1, dressing the seeds.
Furthermore, the mixed matrix comprises the following components in percentage by mass: and (3) sterilized soil: humus soil: organic fertilizer is 2: 1: 1.
Furthermore, the humidity of the mixed matrix is 50% -70% in the seedling hardening process.
Furthermore, the mixed solution A is prepared by mixing an organic water-soluble fertilizer and N: p: k is 20: 20: 20 of mixed liquid of water-soluble foliar fertilizer; wherein the dosage of the organic water-soluble fertilizer is 45 g/mu; the dosage of the water-soluble foliar fertilizer is 60 ml/mu.
Further, the mixed solution B is prepared by mixing an organic water-soluble fertilizer and N: p: k-14: 6: 40 of water-soluble leaf fertilizer mixed liquor; wherein the dosage of the organic water-soluble fertilizer is 60 g/mu; the dosage of the water-soluble foliar fertilizer is 75 ml/mu.
According to the rapid breeding method of the polygonatum sibiricum seedlings, provided by the invention, the tuber tissue culture breeding technology is combined with the seedling hardening technology, so that the problems of long breeding time and low seedling standardization degree are solved, the problems of long seedling time, irregular seedling emergence, seed and seed source mixing and the like in direct seeding breeding of polygonatum sibiricum seeds are solved, and in addition, the selected material for tissue culture is the tuber, so that the pure positive of the seed source can be ensured. Additional features and advantages of the invention will be set forth in the description which follows, and in part will be obvious from the description, or may be learned by practice of the invention. The objectives and other advantages of the invention will be realized and attained by the structure particularly pointed out in the written description and drawings.
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In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the embodiments or the prior art will be briefly described below, and it is obvious that the drawings in the following description are some embodiments of the present invention, and those skilled in the art can also obtain other drawings according to the drawings without creative efforts.
FIG. 1 shows a schematic representation of a tuber after treatment according to an embodiment of the present invention;
FIG. 2 is a schematic view showing the state of primary culture of a tissue culture seedling according to an embodiment of the present invention;
FIG. 3 is a diagram showing the state of subculture of tissue-cultured seedlings according to an embodiment of the present invention;
FIG. 4 is a schematic diagram showing the culture state of proliferation and differentiation of a clumpy bud according to an embodiment of the present invention;
FIG. 5 is a schematic diagram showing the state of rooting culture of differentiated clumpy buds according to an embodiment of the present invention;
FIG. 6 is a diagram illustrating the growth of a seedling before hardening off according to an embodiment of the present invention;
FIG. 7 is a diagram showing the growth of the seedlings after hardening off the tissue culture seedlings for 3 months according to the embodiment of the present invention.
Detailed Description
In order to make the objects, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are some, but not all, embodiments of the present invention. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
In order to solve the problems of long seedling raising time, irregular seedling emergence, seed and seed source mixing and the like in the direct seeding and seedling raising of sealwort seeds, a method combining a rhizome tissue culture breeding technology with a seedling hardening technology is adopted for breeding. At present, many intermediate types of polygonatum belong to the genus are hybrid types caused by concentrated cultivation of various polygonatum. Because the seeds are the products of the pollination of the reproductive organs of the polygonatum kingianum, flowers belonging to the internodes can be pollinated mutually, and the rootstocks are the nutritive organs of the polygonatum kingianum, have the hereditary property of the seeds and can not be hybridized, the seed source can be kept pure; therefore, compared with seed breeding, rhizome breeding can ensure the pureness of seed sources. The breeding method solves the problems that the breeding time of the polygonatum sibiricum direct-seeding seedlings is too long, and the seedling standardization degree is low.
The tissue culture method adopts rootstocks as the selected materials, and adopts the following specific technical measures:
1. determining the collection time of the rhizome of the polygonatum sibiricum tissue culture and the selection conditions of the rhizome;
2. the method comprises determining rhizome tissue culture seedling breeding technique, including sterilization, primary culture, subculture, cluster bud proliferation and differentiation, and rooting culture, wherein the production time of the tissue culture seedling is 6 months;
3. the seedling hardening technology of the tissue culture seedlings is determined, which comprises the selection of a matrix, the application of a seedling hardening seed dressing material and a seedling hardening fertilizer, the seedling hardening of 3 months is ensured to enable the seedlings to adapt to the field growth, and the root and stem diameter of the seedlings is more than or equal to 2.5 cm.
Specifically, the rapid breeding method of the sealwort seedlings comprises the following steps:
1. tissue culture seedling breeding
(1) Selection of rootstocks: selecting different seed sources according to different requirements between 11 months of each year and 2 months of the next year, and digging rhizomes of Polygonatum kingianum of the confirmed seed sources, wherein the seed sources exemplarily comprise Polygonatum kingianum (Polygonatum kingianum Coll. et Hemsl.), Polygonatum kingianum (Polygonatum sibiricum Delar. ex Redoute) and Polygonatum cyrtonema Hua; selecting strong, rot-free, disease and pest-free fresh rhizome of rhizoma Polygonati with bud and multiple bud points, and cutting off dead branches and fibrous roots.
Because many species of polygonatum exist, the polygonatum medicinal material pharmacopoeia has three basic sources, and the prices of different species of seedlings are different, the seed source of the tissue culture raw material needs to be confirmed before tissue culture, and then the seed source of the tissue culture seedling is confirmed.
(2) And (3) sterilizing tubers: washing selected rhizoma Polygonati tubers with sand by using a small brush, and washing the rhizoma Polygonati tubers by using a detergent, wherein the detergent in the application comprises but is not limited to: detergent, washing powder, soap and soap powder, wherein in the embodiment, the washing powder is selected as a detergent, a small amount of the washing powder is used for cleaning the selected rhizoma polygonati tubers for 5-8min, and then the rhizoma polygonati tubers are washed for 2-3h by running water; sterilizing the cleaned rhizoma Polygonati tuber with 75% alcohol for 25-35s, washing with sterile water for 2-3 times, sterilizing with 0.2% mercuric chloride for 10-15min, and cleaning with sterile water for 6-7 times. Tubers after treatment are shown in FIG. 1.
(3) Primary culture of tubers: cutting rhizoma Polygonati with bud and tuber into 1-3cm, and inoculating on a first culture medium; culturing in 25 deg.C culture room under light illumination for 10 hr and dark 14 hr at 6000lux light intensity for 7-14 days to obtain cluster bud. The schematic diagram of the state of the primary culture of the tissue culture seedling is shown in fig. 2.
The first culture medium is a culture medium which takes MS as a basic culture medium and is added with 3.0mg/L BA, 1.0mg/L LNAA, 30g/L sucrose and 5g/L agar.
(4) Subculturing cluster buds: the subculture refers to the transfer of the medium and the recut culture of the clumpy buds after the primary culture of tubers. Removing branches and leaves of the cluster buds, cutting into tubers with the size of 1cm, inoculating the tubers on a second culture medium, placing the tubers on a culture room with the temperature of 25 ℃ for culture, illuminating for 10h and darkness for 14h, and obtaining secondary cluster buds after 20-30 days, wherein the light intensity is 6000 lux. The state diagram of the tissue culture seedling subculture is shown in fig. 3.
Wherein the second culture medium is a culture medium which takes MS as a basic culture medium and is added with 4.0mg/L BA, 0.5mg/L2,4-D, 30g/L sucrose and 5g/L agar.
(5) Clumpy bud proliferation and differentiation: removing branches and leaves of the secondary multiple buds, cutting into tubers with the size of 1cm, inoculating the tubers on a third culture medium, culturing in a culture room with the temperature of 25 ℃, illuminating for 10 hours and darkness for 14 hours, and obtaining value-added multiple buds after 30 days;
Removing branches and leaves of the proliferated multiple buds, cutting into tubers with the size of 1cm, inoculating the tubers on a fourth culture medium, placing the tubers on a culture room with the temperature of 25 ℃ for culture, and illuminating for 10h and dark for 14h at the light intensity of 6000lux, and obtaining differentiated multiple buds after 30 days. A schematic of the culture state of proliferation and differentiation of the multiple shoots is shown in FIG. 4.
Wherein the third culture medium is a culture medium which takes MS as a basic culture medium and is added with 2.0mg/L BA, 0.5mg/L LNAA, 30g/L sucrose and 5g/L agar;
the fourth culture medium is a culture medium which takes MS as a basic culture medium and is added with 1.0mg/L BA, 0.2mg/L NAA, 30g/L sucrose and 5g/L agar.
(6) And (3) rooting culture of differentiated cluster buds: and (3) cutting tubers with buds from the cluster buds into tubers with buds, inoculating the tubers with buds to a fifth culture medium, placing the tubers with buds into a culture room at 25 ℃ for culture, and obtaining tissue culture seedlings with a large number of roots after 60 days, wherein the tubers are illuminated for 10 hours and are dark for 14 hours and the light intensity is 6000 lux. A schematic representation of the rooting culture state of the differentiated clumpy buds is shown in FIG. 5.
Wherein the fifth culture medium is a culture medium which takes 1/2MS as a basic culture medium and is added with 1.0mg/LNAA, 0.5g/L active carbon, 30g/L sucrose and 5g/L agar.
2. Hardening off tissue culture seedling
(1) Hardening seedlings: moving the rooted aseptic seedlings to a greenhouse, carrying out adaptive seedling hardening for 7 days under natural conditions, not opening a culture bottle cap in the seedling hardening process, then taking out the rooted aseptic seedlings, cleaning a culture medium, cleaning seedlings cultured by tissue after the culture medium is cleaned, namely a seedling growth condition schematic diagram before seedling hardening is shown in figure 6, slightly airing, putting the aseptic seedlings cleaned with the culture medium into a container, and carrying out adaptive seedling hardening by using carbendazim: iprodione: thiram is 1: 1: 1, dressing seeds, ensuring that 80-85% of the area of the roots of aseptic seedlings is stained with bactericide mixed powder when the dressing is finished, and transplanting the seedlings to soil with the mass ratio of sterilized soil: humus soil: organic fertilizer is 2: 1: 1, the seedlings begin to turn green after 30 days.
"Regreenness" is a term used in cultivation, which means that after a seedling of a plant is transplanted or overwintering, it changes from yellow to green and growth is restored. Wherein, the standard of striking green in this application is exactly that after the tissue culture seedling is planted, the leaf begins to resume normal green state, grows well, has new gemmule to take place.
(2) Hardening seedling management: after the aseptic seedlings are transplanted to the matrix, spraying the mixed solution A after 40 days; spraying the mixed solution B after 60 days, and hardening seedlings after 90 days to obtain sealwort seedlings.
Specifically, the mixed solution A is an organic water-soluble fertilizer and N: p: k is 20: 20: 20 of mixed liquid of water-soluble foliar fertilizer; wherein the dosage of the organic water-soluble fertilizer is 45 g/mu; the dosage of the water-soluble foliar fertilizer is 60 ml/mu, and the mixed solution B is the mixture of the organic water-soluble fertilizer and N: p: k-14: 6: 40 of water-soluble leaf fertilizer mixed liquor; wherein the dosage of the organic water-soluble fertilizer is 60 g/mu; the dosage of the water-soluble foliar fertilizer is 75 ml/mu.
For example, the organic water-soluble fertilizer of the embodiment is a ganle organic water-soluble fertilizer; after aseptic seedlings are transplanted to a matrix, keeping the humidity of the matrix to be about 60%, turning green the seedlings by more than 80% after 40 days, spraying once Ganle organic water-soluble fertilizer (45 g/mu) and N: p: k is 20: 20: 20, spraying 60 g/mu of organic water-soluble fertilizer (60 g/mu) and N: p: k-14: 6: 40 of water-soluble foliar fertilizer (75 ml/mu) mixed liquor, hardening off the tissue culture seedlings for 3 months, wherein the seedling growth condition schematic diagram after hardening off the tissue culture seedlings for 3 months is shown in figure 7, the statistical survival rate after 3 months is achieved, and the hardening-off survival rate is more than 90%.
The invention combines two fertilizers by selecting and using the mixed solution of the two fertilizers, which is beneficial to improving the survival rate of the tissue culture seedling in the seedling hardening process, and simultaneously can improve the biological yield and promote the rapid growth of the rootstocks.
The rapid breeding method of the sealwort seedlings provided by the application of the invention effectively promotes the growth speed of sealwort, improves the quality of sealwort seedlings, realizes standardized and industrialized production of sealwort seedlings, obtains a large amount of high-quality sealwort seedlings and meets the demand of sealwort planting. The invention provides a polygonatum sibiricum seedling breeding method which can shorten the tissue culture time and improve the seedling quality and the transplanting survival rate by the innovative design of a polygonatum sibiricum tissue culture formula and a breeding method combining rhizome tissue culture and seedling hardening technology.
Illustratively, the breeding example of polygonatum kingianum seedlings is carried out by using the breeding method provided by the application:
(1) selection of rootstocks: and 7 days 12 and 7 months in 2018, digging rhizomes of polygonatum kingianum, selecting fresh polygonatum kingianum rhizomes which are strong in growth, free of decay and diseases and insect pests and have more buds and buds, and shearing off dead branches and fibrous roots.
(2) And (3) sterilizing tubers: washing 2kg of selected polygonatum kingianum tubers with sand and 20g of washing powder for 5min by using a small brush, and washing for 2h by using running water; sterilizing the cleaned rhizoma Polygonati stem tuber with 75% alcohol for 30s, washing with sterile water for 2-3 times, sterilizing with 0.2% mercuric chloride for 12min, and cleaning with sterile water for 6-7 times.
(3) The primary culture, the secondary culture, the propagation and differentiation of cluster buds and the rooting culture of tubers are all cultured according to the requirements and the method, and the tissue culture seedlings are transferred to a greenhouse for hardening seedlings after 165 days.
(4) Hardening seedlings: hardening seedlings for 7 days under natural conditions, hardening the seedlings according to the application, and turning green the seedlings after 35 days; after 44 days, more than 80 percent of seedlings basically have 1 leaf; according to the requirements of the method, foliar fertilizer is sprayed and water management is carried out, seedlings are picked and dug for detection after hardening for 94 days, the hardening survival rate is 96%, the average diameter of roots and stems of 60 seedlings of tissue culture seedlings is 2.71cm, the minimum diameter is 2.14cm, and the maximum diameter is 3.11 cm; the fibrous root is larger than 5, and the bud head is larger than 2. After transplanting the field for 3 months, the survival rate of the field is measured to be 93 percent.
The rapid breeding method of the sealwort seedlings provided by the invention can effectively solve the problems of long seedling growing time, irregular seedling emergence, mixed seed and seed sources and high cost of direct seeding and seedling growing of sealwort seeds, effectively promote the growth speed of sealwort, improve the quality of sealwort seedlings, realize standardized and industrialized production of sealwort seedlings, obtain a large amount of high-quality sealwort seedlings and meet the demand of sealwort planting.
Although the present invention has been described in detail with reference to the foregoing embodiments, it will be understood by those of ordinary skill in the art that: the technical solutions described in the foregoing embodiments may still be modified, or some technical features may be equivalently replaced; and such modifications or substitutions do not depart from the spirit and scope of the corresponding technical solutions of the embodiments of the present invention.
Claims (15)
1. A rapid breeding method of sealwort seedlings is characterized in that: the method comprises the following steps:
1) obtaining sealwort tissue culture tubers;
2) carrying out tissue culture on the sealwort tissue culture tubers by using a tissue culture seedling breeding technology to obtain tissue culture seedlings;
3) and (3) carrying out hardening culture on the tissue culture seedlings by utilizing a tissue culture hardening-seedling technology until the diameter of the hardened-seedling rhizome is more than or equal to 2.5cm, and obtaining the required seedlings.
2. The rapid propagation method of sealwort seedlings according to claim 1, characterized in that: the method for obtaining the sealwort tissue culture tubers comprises the following steps:
obtaining rhizome of polygonatum sibiricum with a confirmed seed source, wherein the rhizome of polygonatum sibiricum is a fresh rhizome of polygonatum sibiricum which is strong in growth, free of rot, free of plant diseases and insect pests and provided with buds and buds, and removing dead branches and fibrous roots to obtain rhizome tubers;
cleaning the obtained rhizoma Polygonati tuber, washing with detergent for 5-8min, and washing with running water for 2-3 h; sterilizing the cleaned rhizoma Polygonati tuber with 75% alcohol for 25-35s, washing with sterile water for 2-3 times, sterilizing with 0.2% mercuric chloride for 10-15min, and cleaning with sterile water for 6-7 times;
cutting rhizoma Polygonati stem with bud into 1-3cm diameter rhizoma Polygonati tissue culture tuber.
3. The rapid propagation method of sealwort seedlings according to claim 1 or 2, characterized in that: the tissue culture cultivation of the sealwort tissue culture tubers by using the tissue culture seedling breeding technology comprises the following steps:
Inoculating the sealwort tissue culture tubers on a first culture medium, and culturing for 7-14 days in a culture room to obtain cluster buds;
removing branches and leaves of the cluster buds, cutting the cluster buds into tubers with the diameter of 1cm, inoculating the tubers on a second culture medium, and culturing in a culture room for 20-30 days to obtain subculture cluster buds;
removing branches and leaves of the subculture multiple buds, cutting into tubers with the diameter of 1cm, inoculating the tubers on a third culture medium, and culturing in a culture room for 30d to obtain proliferation multiple buds;
removing branches and leaves of the proliferated multiple buds, cutting the multiple buds into tubers with the diameter of 1cm, and inoculating the tubers onto a fourth culture medium; culturing in a culture room for 30 days to obtain differentiated cluster buds;
and cutting the cluster buds into tubers with buds, inoculating the tubers with the buds onto a fifth culture medium, and culturing for 60 days in a culture chamber to obtain tissue culture seedlings with roots.
4. The rapid propagation method of sealwort seedlings according to claim 3, characterized in that: the culture conditions of the culture chamber are as follows: at 25 ℃, the light is 10h, the dark is 14h, and the light intensity is 6000 lux.
5. The rapid propagation method of sealwort seedlings according to claim 4, characterized in that: the first culture medium is: MS is taken as a basic culture medium, and a culture medium of 3.0mg/LBA, 1.0mg/LNAA, 30g/L sucrose and 5g/L agar is added.
6. The rapid propagation method of sealwort seedlings according to claim 4, characterized in that: the second culture medium is: MS is taken as a basic culture medium, and a culture medium of 4.0mg/L BA, 0.5 mg/L2, 4-D, 30g/L sucrose and 5g/L agar is added.
7. The rapid propagation method of sealwort seedlings according to claim 4, characterized in that: the third culture medium is: MS is taken as a basic culture medium, and a culture medium of 2.0mg/LBA, 0.5mg/LNAA, 30g/L sucrose and 5g/L agar is added.
8. The rapid propagation method of sealwort seedlings according to claim 4, characterized in that: the fourth culture medium is: MS is taken as a basic culture medium, and a culture medium of 1.0mg/LBA, 0.2mg/LNAA, 30g/L sucrose and 5g/L agar is added.
9. The rapid propagation method of sealwort seedlings according to claim 4, characterized in that: the fifth culture medium is: 1/2MS is used as a basic culture medium, and a culture medium of 1.0mg/LNAA, 0.5g/L active carbon, 30g/L sucrose and 5g/L agar is added.
10. The rapid propagation method of sealwort seedlings according to claim 1 or 2, characterized in that: the tissue culture seedling exercising culture by utilizing the tissue culture seedling exercising technology comprises the following steps:
taking out the rooted aseptic seedlings, dressing the seedlings, transplanting the seedlings into a mixed matrix, and turning green the seedlings after 30 days;
spraying the mixed solution A after the aseptic seedlings are transplanted to the matrix for 40 days; spraying the mixed solution B after 60 days, and hardening seedlings after 90 days to obtain sealwort seedlings.
11. The rapid propagation method of polygonatum sibiricum seedlings according to claim 10, wherein: the seed dressing comprises the following steps: putting the sterile seedlings of the clean culture medium into a suitable container, and adding the mixture of the components in percentage by mass as follows: iprodione: thiram is 1: 1: 1, dressing the seeds.
12. The rapid propagation method of polygonatum sibiricum seedlings according to claim 10, wherein: the mixed matrix comprises the following components in percentage by mass: and (3) sterilized soil: humus soil: organic fertilizer is 2: 1: 1.
13. The rapid propagation method of polygonatum sibiricum seedlings according to claim 10, wherein: in the seedling exercising process, the humidity of the mixed matrix is 50% -70%.
14. The rapid propagation method of polygonatum sibiricum seedlings according to claim 10, wherein: the mixed liquid A is organic water-soluble fertilizer and N: p: k is 20: 20: 20 of mixed liquid of water-soluble foliar fertilizer; wherein the dosage of the organic water-soluble fertilizer is 45 g/mu; the dosage of the water-soluble foliar fertilizer is 60 ml/mu.
15. The rapid propagation method of polygonatum sibiricum seedlings according to claim 10, wherein: the mixed solution B is organic water-soluble fertilizer and N: p: k-14: 6: 40 of water-soluble leaf fertilizer mixed liquor; wherein the dosage of the organic water-soluble fertilizer is 60 g/mu; the dosage of the water-soluble foliar fertilizer is 75 ml/mu.
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