CN112106655A - Tissue culture method of plateau prunus mume - Google Patents
Tissue culture method of plateau prunus mume Download PDFInfo
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- CN112106655A CN112106655A CN201910530478.8A CN201910530478A CN112106655A CN 112106655 A CN112106655 A CN 112106655A CN 201910530478 A CN201910530478 A CN 201910530478A CN 112106655 A CN112106655 A CN 112106655A
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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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- 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 discloses a tissue culture method of plateau prunus mume, which comprises the following steps: and (3) seedling cultivation: selecting seedlings: selecting a wild cherry plum seedling with the flowering amount, the fruit bearing rate and the average fruit diameter all being close to the front in the third year, selecting a terminal bud as an explant and sterilizing the explant when the wild cherry plum seedling sprouts next time, and shearing the terminal bud according to the number of leaves on the terminal bud; C. culturing: c1, inoculating the explant to the primary culture medium, culturing for 2-3 generations, and transplanting the explant to the proliferation culture medium; c2, rejuvenating the seedlings in 70 th to 100 th days of culture; c3, transferring the rejuvenated seedlings to a rooting culture medium; D. hardening seedlings: and (4) hardening the rooted seedlings with developed root systems in a tissue culture room, and then transferring the rooted seedlings to a greenhouse for hardening the seedlings until the seedlings grow to 18-24 centimeters. The method can not only improve the propagation speed, but also greatly improve the drought resistance, frost resistance, high fruiting rate and other characteristics of the seedlings after tissue culture, keep the short characteristics of the Prunus serrulata, and greatly improve the survival rate.
Description
Technical Field
The invention relates to the field of plant tissue culture methods, in particular to a tissue culture method of plateau prunus mume.
Background
The wild cherry berries are commonly called as the aronia melanocarpa, integrate the values of eating, medicine, garden, ecology and the like, can be eaten raw, can also be made into wine, jam, syrup, fruit juice, tea, salad, beer and ice cream, contain rich vitamins, minerals, antioxidants and other important polyphenols, and have the effect of resisting cancer cells.
Wild cherry berries are native to wet forests and swamps in North America and introduced into Europe and east Asia, and in the nineties of the last century, wild cherry berries were introduced into China and planted in the western Liaoning. At present, the wild cherry berries are planted in a plurality of provinces of China. However, the existing aronia serrulata L are planted in the plateau area of hills, and in the plateau area, such as the Tibet area, the climate is severe, the average altitude is more than 4000 meters, and the plants are not easy to survive. The economic development of the existing Tibetan area is extremely unbalanced, poverty relief is an important thing for poverty-stricken areas, wild cherry berries have high commercial value, the value of the existing wild cherry berry extract is about several hundreds to one thousand, the existing wild cherry berry extract is one of excellent items for poverty relief, tissue culture is a plant which can be used for rapidly propagating some rare plants or has high economic value, how to apply the tissue culture to the planting of the wild cherry berries makes the tissue culture meet the requirements of rapid large-area planting, and the performance stability is a thing for research.
Disclosure of Invention
The invention provides a tissue culture method of plateau prunus mume in order to solve the technical problems.
The invention is realized by the following technical scheme:
a tissue culture method of plateau prunus mume comprises the following steps:
A. and (3) seedling cultivation:
a1, selecting a planting area, deep ploughing and turning soil in the area, and improving the soil in the area by adopting 55-65% of bio-organic fertilizer and 35-45% of bio-organic bacteria by weight to ensure that the pH value of the soil is 6.6-7; planting the prunus laevigata after soil improvement for 4 months;
a2, applying organic fertilizer to the Prunus serrulata seedlings in the first flowering phase;
a3, pruning the seedlings of the Prunus serrulata after the first fruit harvest, and burying cake fertilizer beside the seedlings;
B. selecting seedlings: selecting a wild cherry plum seedling with the flowering amount, the fruit bearing rate and the average fruit diameter all being close to the front in the third year, selecting a terminal bud as an explant and sterilizing the explant when the wild cherry plum seedling sprouts next time, and shearing the terminal bud according to the number of leaves on the terminal bud;
C. culturing:
c1, inoculating the explant to the primary culture medium, culturing for 2-3 generations, and transplanting the explant to the proliferation culture medium;
c2, rejuvenating the seedlings in 70 th to 100 th days of culture;
c3, transferring the rejuvenated seedlings to a rooting culture medium;
D. hardening seedlings: and (4) hardening the rooted seedlings with developed root systems in a tissue culture room, and then transferring the rooted seedlings to a greenhouse for hardening the seedlings until the seedlings grow to 18-24 centimeters.
The plateau area condition is abominable, is influenced by environment and weather and hardly relies on natural condition fast and breeds large batch prunus mume, and the prunus mume can be along with weather, environmental variation to the regional environment is planted in the adaptation, and only the prunus mume that adapts to the plateau environment can keep the fruit bearing rate. Selecting excellent seedlings in one region for the tissue cultured seedlings so as to keep the excellent adaptogen habits of the tissue cultured seedlings of the Prunus serrulata. Soil is improved to be weakly acidic before planting, organic fertilizer is applied in the first flowering phase, cake fertilizer is applied after fruits are harvested, and the fruit quality is effectively improved. The soil quality of plateau areas such as Tibet areas is different from that of inland plain areas, and before planting, soil improvement is needed to be carried out on the plateau areas so as to meet the growth requirement of the lantana camara. The cake fertilizer is residue left after oil extraction of oil seeds, contains a large amount of nutrient elements, and the cake is a slow-release fertilizer, adopts a shallow burying mode, avoids directly fertilizing the prunus mume, and can be fermented and matured firstly when being buried shallowly, so that the damage to the root system is reduced, and the growth and development of the prunus mume are improved. Adopt 18 to 24 outdoor planting of seedling transplantation behind the plant tissue culture, can avoid wild cherry plum tree to transplant the highland after the variation grow excessively, keep the short and small characteristic of wild cherry plum tree, can improve the survival rate greatly, the wild cherry plum tree height of planting of this scheme of adoption is between 1 meter to 1.5 meters, more is suitable for the plateau to plant. In the scheme, the terminal buds of the Prunus serrulata branches with high fruiting rate in the region are used as explants, so that the propagation speed can be increased, and the characteristics of drought resistance, frost resistance, high fruiting rate and the like of seedlings after tissue culture can be greatly improved.
Preferably, in step B, the number of leaves of the terminal bud is at least 3, and the number of the cut leaves is equal to the number of the leaves.
Preferably, the biological organic bacteria adopt the following strains and parts by weight: 20-30 parts of bacillus subtilis, 10-15 parts of bacillus laterosporus and 25-28 parts of EM (effective microorganisms). Plateau soil is infertile, adopt bio-organic fertilizer + bio-organic fungus, need carry out high temperature sterilization to it and handle, general bacterial disinfects under 60 degrees of temperature, but, because plateau district bio-organic fertilizer's generally come from livestock such as ox, sheep, it needs to disinfect a week at the temperature more than 90 degrees centigrade, go on in order to guarantee the stability of its follow-up fermentation, this embodiment adopts above-mentioned 3 bacterial, can guarantee still can keep active in a week more than 90 degrees, also can improve plateau soil well, improve the fruit quality of producing of prunus mume.
Preferably, the biological organic bacteria adopt the following strains and parts by weight: 24 parts of bacillus subtilis, 11 parts of bacillus laterosporus and 258 parts of EM bacteria.
Furthermore, 60% by weight of bio-organic fertilizer and 40% by weight of bio-organic bacteria are used for improving the soil.
Preferably, the proliferation medium is 0.065-0.78mg/L of forchlorfenuron, 20-25g/L of sucrose, 6-7g/L of agar, 0.025-0.03 g/L of naphthylacetic acid and 0.01-0.02 g/L of gibberellin.
Preferably, the proliferation medium is 0.7mg/L of forchlorfenuron, 25g/L of sucrose, 6g/L of agar, 0.028 g/L of naphthylacetic acid and 0.015 g/L of gibberellin.
Compared with the prior art, the invention has the following advantages and beneficial effects:
1. the invention adopts the terminal bud of the Prunus serrulata branch with high fruiting rate in the region as the explant, which can not only improve the propagation speed, but also greatly improve the drought resistance, frost resistance, high fruiting rate and other characteristics of the nursery stock after tissue culture.
The outdoor transplantation of 18-24 seedlings after the tissue culture of the plants can avoid the excessive variation of the wild cherry plum trees after the transplantation of plateau, keep the short and small characteristics of the wild cherry plum trees, and greatly improve the survival rate which can reach 99 percent.
Detailed Description
In order to make the objects, technical solutions and advantages of the present invention more apparent, the present invention is further described in detail below with reference to examples, and the exemplary embodiments and descriptions thereof are only used for explaining the present invention and are not used as limitations of the present invention.
Example 1
A tissue culture method of plateau prunus mume comprises the following steps:
A. and (3) seedling cultivation:
a1, selecting a planting area and deeply ploughing and turning soil in the area to improve the soil looseness, so as to provide enough oxygen for the root system respiration after the subsequent planting of the cerasus serrulata, and promote the root system development; improving the soil in the area by adopting 55-65% of bio-organic fertilizer and 35-45% of bio-organic bacteria by weight ratio to ensure that the pH value of the soil is 6.6-7; planting the prunus laevigata after soil improvement for 4 months;
a2, applying organic fertilizer to the Prunus serrulata seedlings in the first flowering phase;
a3, pruning the seedlings of the Prunus serrulata after the first fruit harvest, and burying cake fertilizer beside the seedlings;
B. selecting seedlings: selecting a wild cherry plum seedling with the flowering amount, the fruit bearing rate and the average fruit diameter all being close to the front in the third year, selecting a terminal bud as an explant and sterilizing the explant when the wild cherry plum seedling sprouts next time, and shearing the terminal bud according to the number of leaves on the terminal bud;
C. culturing:
c1, inoculating the explant to the primary culture medium, culturing for 2-3 generations, and transplanting the explant to the proliferation culture medium;
c2, rejuvenating the seedlings in 70 th to 100 th days of culture;
c3, transferring the rejuvenated seedlings to a rooting culture medium;
D. hardening seedlings: and (4) hardening the rooted seedlings with developed root systems in a tissue culture room, and then transferring the rooted seedlings to a greenhouse for hardening the seedlings until the seedlings grow to 18-24 centimeters.
The soil of plateau areas such as Tibet areas is barren, different from the soil quality of inland soil, and the soil of the areas is improved firstly when seedlings are cultivated, so that the soil virtuous degree is improved, and the growth requirement of the Prunus serrulata is met. The soil is improved by adopting the combination of the biological organic fertilizer and the biological organic bacteria so as to improve the virtuous degree of the soil. The prunus laevigata bred by the tissue culture method is suitable for plateau environment planting, has high survival rate, can be planted in a plurality of regions in Tibet, has the survival rate of over 98 percent, high fruit bearing rate and full fruits, and has the anthocyanin content which is at least 8 times that of the prunus laevigata in inland.
Example 2
The present embodiment is optimized on the basis of the above embodiments, that is, specifically:
in the implementation land of the embodiment, one land is selected as a seedling cultivation area in a region with the elevation of about 3000 meters in Tibet, particularly around eight-Hotel county, and the area of the region is 10 mu. Deep ploughing and turning soil in the area, wherein the deep ploughing depth is 10 cm-20 cm, so that the soil looseness is improved, sufficient oxygen is provided for root respiration after the subsequent wild cherry plum is planted, and the development of the root system is promoted; and (3) improving the soil in the area by adopting 60% of bio-organic fertilizer and 40% of bio-organic bacteria by weight to ensure that the pH value of the soil is 6.7. The biological organic bacteria adopt the following strains and parts by weight: 20-30 parts of bacillus subtilis, 10-15 parts of bacillus laterosporus and 25-28 parts of EM (effective microorganisms), and specifically, 24 parts of bacillus subtilis, 11 parts of bacillus laterosporus and 258 parts of EM (effective microorganisms) are adopted in the seedling cultivation area. The biological organic fertilizer is prepared from livestock manure. The wild cherry plum is planted 4 months after the soil improvement operation is completed, and the wild cherry plum adopts seedlings cultivated by the afforestation research institute in the arid area of Liaoning province.
Applying organic fertilizer to the wild cherry seedlings in the first flowering period,
pruning the seedlings of the Prunus serrulata after the first fruit harvest, namely approximately planting the Prunus serrulata in the next year, and burying cake fertilizer beside the seedlings, wherein the cake fertilizer is buried shallowly in a manner of being 12-16 cm away from the Prunus serrulata and 5cm deep to realize shallow burying. The cake fertilizer is rapeseed cake fertilizer, can be directly used, and can also be applied after fermentation and decomposition. When the fermented and rotten vegetable seeds are fermented and matured, the rapeseed cakes are added into the livestock manure according to the mass ratio of 1:1, the manure is sprayed with water, mixed and piled up, and sealed by a membrane, and when the membrane is inflated, the side of the membrane is buried beside the wild cherry nursery stocks, and the fertilizing amount per mu is 120 jin.
Selecting a wild cherry seedling with the earlier flowering amount, fruit bearing rate and average fruit diameter in the third year, wherein at least 2 indexes of the flowering amount, fruit bearing rate and average fruit diameter of the wild cherry seedling in a plurality of seedlings are optimal. When the wild cherry plum seedlings sprout next time, selecting 1-2 cm apical buds with at least 3 leaves on the branches as explants and disinfecting the explants, and shearing the explants according to the number of the leaves on the apical buds, wherein the shearing number is equal to the corresponding number of the leaves.
After cutting the terminal bud, inoculating the terminal bud to a primary culture medium, culturing for 2-3 generations, and transplanting the terminal bud to a proliferation culture medium. The primary culture medium can adopt the existing WPM culture medium. The embodiment adopts the value-added culture medium which comprises 0.065-0.78mg/L of forchlorfenuron, 20-25g/L of cane sugar, 6-7g/L of agar, 0.025-0.03 g/L of naphthylacetic acid and 0.01-0.02 g/L of gibberellin. In the range of the mixture ratio, the inventor prepares a plurality of groups of proliferation culture media, and the specific formula is as follows:
ingredient of Pilcolone sucrose agar gibberellin naphthylacetic acid
Group 10.65 mg/L20 g/L6 g/L0.025 g/L0.01 g/L
Group 20.68 mg/L22 g/L6.5 g/L0.026 g/L0.07 g/L
Group 30.7 mg/L25 g/L6 g/L0.028 g/L0.015 g/L
Group 40.74 mg/L22 g/L6.4 g/L0.028 g/L0.01 g/L
Group 50.78 mg/L24 g/L7 g/L0.03 g/L0.02 g/L
The 5 groups of propagation mediums are adopted to perform propagation culture on the 5 groups of terminal buds respectively, wherein the propagation speed is high, but the propagation speed of the group 3 is the fastest.
Rejuvenating the seedlings in 70 th to 100 th days of culture; and transferring the rejuvenated seedlings to a rooting culture medium for rooting.
And (4) hardening the rooted seedlings with developed root systems in a tissue culture room, and then transferring the rooted seedlings to a greenhouse for hardening the seedlings until the seedlings grow to 18-24 centimeters. Preferably, 20 centimeters of nursery stocks are selected for cultivation.
The tissue culture technology of this scheme of adoption can realize the rapid propagation of plateau wild cherry plum, provides effective guarantee for people's appreciation income-creating of Tibetan region, and the wild cherry plum nursery stock after the tissue culture has kept the good gene that adapts to the plateau environment, the bearing rate is high, the anthocyanin content is high, avoids wild cherry plum to vary etc. in the planting process. The cultivation method in the scheme is also preferably selected for the wild cherry seedlings after tissue culture.
The above-mentioned embodiments are intended to illustrate the objects, technical solutions and advantages of the present invention in further detail, and it should be understood that the above-mentioned embodiments are merely exemplary embodiments of the present invention, and are not intended to limit the scope of the present invention, and any modifications, equivalent substitutions, improvements and the like made within the spirit and principle of the present invention should be included in the scope of the present invention.
Claims (7)
1. A tissue culture method of plateau prunus mume is characterized by comprising the following steps:
A. and (3) seedling cultivation:
a1, selecting a planting area, deep ploughing and turning soil in the area, and improving the soil in the area by adopting 55-65% of bio-organic fertilizer and 35-45% of bio-organic bacteria by weight to ensure that the pH value of the soil is 6.6-7; planting the prunus laevigata after soil improvement for 4 months;
a2, applying organic fertilizer to the Prunus serrulata seedlings in the first flowering phase;
a3, pruning the seedlings of the Prunus serrulata after the first fruit harvest, and burying cake fertilizer beside the seedlings;
B. selecting seedlings: selecting a wild cherry plum seedling with the flowering amount, the fruit bearing rate and the average fruit diameter all being close to the front in the third year, selecting a terminal bud as an explant and sterilizing the explant when the wild cherry plum seedling sprouts next time, and shearing the terminal bud according to the number of leaves on the terminal bud;
C. culturing:
c1, inoculating the explant to the primary culture medium, culturing for 2-3 generations, and transplanting the explant to the proliferation culture medium;
c2, rejuvenating the seedlings in 70 th to 100 th days of culture;
c3, transferring the rejuvenated seedlings to a rooting culture medium;
D. hardening seedlings: and (4) hardening the rooted seedlings with developed root systems in a tissue culture room, and then transferring the rooted seedlings to a greenhouse for hardening the seedlings until the seedlings grow to 18-24 centimeters.
2. The tissue culture method of plateau prunus mume as claimed in claim 1, wherein in step B, the number of leaves of the apical bud is at least 3, and the number of the cut leaves is equal to the number of the leaves.
3. The tissue culture method of the plateau prunus mume as claimed in claim 1, wherein the biological organic bacteria adopts the following strains and weight portions: 20-30 parts of bacillus subtilis, 10-15 parts of bacillus laterosporus and 25-28 parts of EM (effective microorganisms).
4. The tissue culture method of the plateau prunus mume as claimed in claim 1, wherein the biological organic bacteria adopts the following strains and weight portions: 24 parts of bacillus subtilis, 11 parts of bacillus laterosporus and 258 parts of EM bacteria.
5. The tissue culture method of plateau prunus mume as claimed in any one of claims 1, 3 and 4, wherein the soil is improved by using 60% by weight of bio-organic fertilizer and 40% by weight of bio-organic bacteria.
6. The tissue culture method of plateau prunus mume as claimed in claim 1, wherein the proliferation medium is Piperazone 0.065-0.78mg/L, sucrose 20-25g/L, agar 6-7g/L, naphthylacetic acid 0.025-0.03 g/L, gibberellin 0.01-0.02 g/L.
7. The tissue culture method of plateau prunus mume as claimed in claim 1, wherein the proliferation medium is Piperazone 0.7mg/L, sucrose 25g/L, agar 6g/L, naphthylacetic acid 0.028 g/L, gibberellin 0.015 g/L.
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN101103702A (en) * | 2007-08-24 | 2008-01-16 | 东北林业大学 | Excised reproduction method for mountain ash |
| CN107493905A (en) * | 2017-09-12 | 2017-12-22 | 芜湖市龙潭园景工程有限公司 | A kind of ornamental plant planting technology |
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- 2019-06-19 CN CN201910530478.8A patent/CN112106655A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101103702A (en) * | 2007-08-24 | 2008-01-16 | 东北林业大学 | Excised reproduction method for mountain ash |
| CN107493905A (en) * | 2017-09-12 | 2017-12-22 | 芜湖市龙潭园景工程有限公司 | A kind of ornamental plant planting technology |
Non-Patent Citations (2)
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| 刘长红: "黑果腺肋花楸组培育苗关键技术", 《现代农业科技》 * |
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