CN113173818B - Biological bacterial fertilizer and method for promoting growth of tissue culture seedlings of alpine rhododendron - Google Patents

Biological bacterial fertilizer and method for promoting growth of tissue culture seedlings of alpine rhododendron Download PDF

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CN113173818B
CN113173818B CN202110446433.XA CN202110446433A CN113173818B CN 113173818 B CN113173818 B CN 113173818B CN 202110446433 A CN202110446433 A CN 202110446433A CN 113173818 B CN113173818 B CN 113173818B
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rhododendron
biological bacterial
growth
tissue culture
biological
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CN113173818A (en
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龚记熠
刘杰
李菲
唐婧
唐明
张习敏
乙引
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Guizhou Education University
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    • C—CHEMISTRY; METALLURGY
    • C05—FERTILISERS; MANUFACTURE THEREOF
    • C05G—MIXTURES OF FERTILISERS COVERED INDIVIDUALLY BY DIFFERENT SUBCLASSES OF CLASS C05; MIXTURES OF ONE OR MORE FERTILISERS WITH MATERIALS NOT HAVING A SPECIFIC FERTILISING ACTIVITY, e.g. PESTICIDES, SOIL-CONDITIONERS, WETTING AGENTS; FERTILISERS CHARACTERISED BY THEIR FORM
    • C05G3/00—Mixtures of one or more fertilisers with additives not having a specially fertilising activity
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01C—PLANTING; SOWING; FERTILISING
    • A01C21/00—Methods of fertilising, sowing or planting
    • A01C21/005—Following a specific plan, e.g. pattern
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
    • A01N63/20—Bacteria; Substances produced thereby or obtained therefrom
    • A01N63/22—Bacillus
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
    • A01N63/30—Microbial fungi; Substances produced thereby or obtained therefrom
    • A01N63/32—Yeast
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
    • A01N63/30—Microbial fungi; Substances produced thereby or obtained therefrom
    • A01N63/38—Trichoderma
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01P—BIOCIDAL, PEST REPELLANT, PEST ATTRACTANT OR PLANT GROWTH REGULATORY ACTIVITY OF CHEMICAL COMPOUNDS OR PREPARATIONS
    • A01P21/00—Plant growth regulators
    • C—CHEMISTRY; METALLURGY
    • C05—FERTILISERS; MANUFACTURE THEREOF
    • C05F—ORGANIC FERTILISERS NOT COVERED BY SUBCLASSES C05B, C05C, e.g. FERTILISERS FROM WASTE OR REFUSE
    • C05F11/00—Other organic fertilisers
    • C05F11/08—Organic fertilisers containing added bacterial cultures, mycelia or the like
    • C—CHEMISTRY; METALLURGY
    • C05—FERTILISERS; MANUFACTURE THEREOF
    • C05G—MIXTURES OF FERTILISERS COVERED INDIVIDUALLY BY DIFFERENT SUBCLASSES OF CLASS C05; MIXTURES OF ONE OR MORE FERTILISERS WITH MATERIALS NOT HAVING A SPECIFIC FERTILISING ACTIVITY, e.g. PESTICIDES, SOIL-CONDITIONERS, WETTING AGENTS; FERTILISERS CHARACTERISED BY THEIR FORM
    • C05G3/00—Mixtures of one or more fertilisers with additives not having a specially fertilising activity
    • C05G3/60—Biocides or preservatives, e.g. disinfectants, pesticides or herbicides; Pest repellants or attractants
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    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/14—Fungi; Culture media therefor
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    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/14—Fungi; Culture media therefor
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    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/14—Fungi; Culture media therefor
    • C12N1/16—Yeasts; Culture media therefor
    • C12N1/18—Baker's yeast; Brewer's yeast
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    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/20—Bacteria; Culture media therefor
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    • C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
    • C12R2001/07—Bacillus
    • C12R2001/12—Bacillus polymyxa ; Paenibacillus polymyxa
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    • C12R2001/79—Paecilomyces
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    • C12R2001/00—Microorganisms ; Processes using microorganisms
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    • C12R2001/85—Saccharomyces
    • C12R2001/865—Saccharomyces cerevisiae
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    • C12R2001/885—Trichoderma

Abstract

The invention provides a biological bacterial fertilizer and a method for promoting growth of tissue culture seedlings of alpine rhododendron, and belongs to the technical field of biological bacterial fertilizers. The biological bacterial fertilizer for promoting the growth of the tissue culture seedlings of rhododendron lapponicum comprises saccharomyces cerevisiae, trichoderma longibrachiatum, paecilomyces lilacinus, bacillus polymyxa and pumpkin. According to the invention, through reasonable compounding of the microbial agent and screening of suitable nutrient substances, the biological bacterial fertilizer capable of remarkably improving the survival rate and the seedling strengthening effect of the transplanted tissue culture seedlings of alpine rhododendron is prepared, and high-quality development of the alpine rhododendron industry in China is promoted.

Description

Biological bacterial fertilizer and method for promoting growth of tissue culture seedlings of alpine rhododendron
Technical Field
The invention belongs to the technical field of biological bacterial fertilizers, and particularly relates to a biological bacterial fertilizer and a method for promoting growth of tissue culture seedlings of alpine rhododendron.
Background
Azalea essence is called 'flower and western application', and occupies an important position in the flower market all over the world. The alpine rhododendron has bright color and rich color, and the flowers are large, so that the alpine rhododendron becomes a fine product in the rhododendron. China is a big country of rhododendron resources, and rhododendron lapponicum resources are distributed and concentrated greatly. However, the development of the alpine rhododendron industry is not satisfactory at present, the high-quality development of the alpine rhododendron industry in China is restricted by the defects of an artificial resource cultivation technology, and a technical bottleneck always exists in how to improve the survival rate of tissue culture seedlings and strengthen the seedlings. At present, aiming at the survival rate and the strong seedlings of rhododendron lapponicum tissue culture seedlings, the following modes are adopted, firstly, the management of illumination, temperature, moisture and nutrients in the tissue culture seedling hardening process is optimized, and the environment conditions beneficial to the growth of the tissue culture seedlings are constructed manually, although the mode can improve the survival rate to a certain degree, the control cost of the conditions is higher, the large-area popularization and production are difficult, and meanwhile, the mode has the defect of unsustainable promotion effect on the growth of the seedlings in the later period; and secondly, by optimizing a fertilization mode, the survival rate of tissue culture seedlings and strong seedlings are improved by using inorganic fertilizers, farmyard manures and the like in the forms of matrix fertilizer mixing, root top dressing, foliar fertilization and the like, but the effect is poor, and the defect that the fertilizer effect cannot be sustained for a long time after fertilization exists. Therefore, a method for promoting the growth of the rhododendron lapponicum tissue culture seedling is urgently needed.
Disclosure of Invention
In order to solve the problems, the invention provides a biological bacterial fertilizer and a method for promoting the growth of tissue culture seedlings of rhododendron lapponicum. The biological bacterial fertilizer for promoting the growth of the tissue culture seedlings of alpine rhododendron provided by the invention can obviously improve the survival rate and the seedling strengthening effect of the transplanted tissue culture seedlings of alpine rhododendron and promote the high-quality development of alpine rhododendron industry.
In order to achieve the above purpose, the invention provides the following technical scheme:
the invention provides a biological bacterial fertilizer for promoting the growth of tissue culture seedlings of rhododendron lapponicum, which comprises saccharomyces cerevisiae, trichoderma longibrachiatum, paecilomyces lilacinus, bacillus polymyxa and pumpkin.
Preferably, the biological bacterial fertilizer comprises the following components in parts by volume: 10-15 parts of a saccharomyces cerevisiae bacterial solution, 5-10 parts of a trichoderma longibrachiatum bacterial solution, 10-15 parts of a paecilomyces lilacinus bacterial solution, 15-25 parts of a bacillus polymyxa bacterial solution and 35-60 parts of pumpkin juice;
the viable count of the saccharomyces cerevisiae bacterial liquid is more than or equal to 6 multiplied by 107CFU/mL; the viable count of the trichoderma longibrachiatum bacterial liquid is more than or equal to 4 multiplied by 107CFU/mL; the light purple pseudo-blueThe viable count of the fungus liquid of the mould is more than or equal to 6 multiplied by 107CFU/mL; the viable count of the bacillus polymyxa liquid is more than or equal to 5 multiplied by 108CFU/mL。
Preferably, the culture conditions of the saccharomyces cerevisiae are as follows: the temperature is 27-30 ℃, the time is 24-36 h, and the pH is 4-6.
Preferably, the culture conditions of the trichoderma longibrachiatum are as follows: the temperature is 26-29 ℃, the time is 60-72 hours, and the pH is 4-6.5.
Preferably, the culture conditions of the paecilomyces lilacinus are as follows: the temperature is 27-31 ℃, the time is 36-50 h, and the pH is 4.8-6.2.
Preferably, the culture conditions of the bacillus polymyxa are as follows: the temperature is 28-33 ℃, the time is 30-58 h, and the pH is 6.6-7.8.
The invention provides a method for promoting the growth of tissue culture seedlings of alpine rhododendrons, which applies the biological bacterial manure in the technical scheme to the tissue culture seedlings of alpine rhododendrons.
Preferably, the biological bacterial manure is mixed with other fertilizers or used independently.
Preferably, the biological bacterial fertilizer is used before the alpine rhododendron is transplanted, and is applied for 1 time every 3-4 months after the transplantation; the application amount of the biological bacterial fertilizer is 100-200 mL of the biological bacterial fertilizer applied to every 5kg of the substrate.
Preferably, the biological bacterial manure is revived before use; the resuscitation conditions are as follows: preheating the biological bacterial fertilizer in water at 10-20 ℃ for 60-90 min, and then transferring the biological bacterial fertilizer into water at 20-30 ℃ for placing for 120-180 min.
Advantageous effects
The invention provides a biological bacterial fertilizer for promoting the growth of tissue culture seedlings of rhododendron lapponicum, which comprises saccharomyces cerevisiae, trichoderma longibrachiatum, paecilomyces lilacinus, bacillus polymyxa and pumpkin. The saccharomyces cerevisiae can ferment and decompose carbohydrates around the root system of rhododendron lapponicum and decompose macromolecular carbohydrates into small molecules for the rhododendron lapponicum to absorb. The trichoderma longibrachiatum has the effect of preventing and controlling pathogenic bacteria in the growth process of rhododendron lapponicum; the trichoderma longibrachiatum can generate active metabolites such as chitinase, glucanase, cellulase, protease and the like and damage cell walls of plant pathogenic fungi; but also can secrete antibacterial protein or lyase to inhibit the infection of rhododendron lapponicum pathogenic bacteria; meanwhile, trichoderma longibrachiatum can also secrete extracellular enzymes such as glucosidase and the like to degrade antibiotic toxins generated by pathogenic bacteria and reduce the damage of rhododendron alpinum pathogenic bacteria to beneficial microorganisms of plant root systems. Paecilomyces lilacinus belongs to endoparasitic fungi, has parasitism on pest eggs such as root-knot nematodes, cyst nematodes and the like in rhizosphere soil of rhododendron lapponicum, and acts together with chitin and chitinase secreted by the parasite eggs so as to kill pests; meanwhile, the paecilomyces lilacinus can also generate chemical substances similar to indoleacetic acid to promote the growth and development of root systems, promotes the dissolution of insoluble phosphate through secreted organic acid, is convenient for the root systems of rhododendron lapponicum to absorb, and plays roles in preventing and controlling insect pests and strengthening seedlings. The bacillus polymyxa not only has the biological nitrogen fixation, phosphorus dissolution and potassium dissolution effects, so as to promote the rhododendron lapponicum to absorb nitrogen, phosphorus and potassium in soil; meanwhile, the rhododendron mariae extract can secrete metabolites such as polymyxin and the like to play an antagonistic role on pathogenic microorganisms in soil and plants, and has a good inhibition effect on root rot and leaf spot of rhododendron lapponicum. According to the invention, saccharomyces cerevisiae, trichoderma longibrachiatum, paecilomyces lilacinus and bacillus polymyxa are compounded and acted together, so that the resistance of rhododendron alpinum to pathogenic bacteria and insect pests is obviously improved, and the survival rate of tissue culture seedlings is improved; meanwhile, the absorption of nutrient substances by alpine rhododendron is promoted, and the seedling strengthening effect is good. The pumpkin can provide early-stage nutrient substances for the rapid growth of microorganisms, shortens the recovery and growth time of the microorganisms when the biological bacterial manure is used, and has the characteristics of low cost, easy acquisition, environmental friendliness and the like. According to the invention, through reasonable compounding of the microbial agent and screening of suitable nutrient substances, the biological bacterial fertilizer capable of remarkably improving the survival rate and the seedling strengthening effect of the transplanted tissue culture seedlings of alpine rhododendron is prepared, and the high-quality development of the alpine rhododendron industry in China is promoted; meanwhile, the invention utilizes the self-reproduction and metabolism characteristics of the microorganisms, exerts sustainable action and environment-friendly characteristics, ensures that the fertilizer is effective for a long time after being applied for one or more times, and reduces the cost.
Drawings
FIG. 1 shows the effect of different bacterial manure formulas on the growth of tissue culture seedlings of rhododendron pulchrum;
FIG. 2 is the effect of the biological bacterial manure on the growth of the tissue culture seedlings of Rhododendron delavayi Franch;
FIG. 3 shows the effect of the biological bacterial manure on the growth of the tissue culture seedling of rhododendron delavayi Franch;
FIG. 4 is a diagram showing the growth of the biological bacterial manure of example 1 of the present invention 30 days after the transplantation of Rhododendron nonagromaculatum in a large area use;
FIG. 5 shows the development effect of the root system of Rhododendron pulchrum after the biological bacterial manure treatment.
Detailed Description
The invention provides a biological bacterial fertilizer for promoting the growth of tissue culture seedlings of rhododendron lapponicum, which comprises saccharomyces cerevisiae, trichoderma longibrachiatum, paecilomyces lilacinus, bacillus polymyxa and pumpkin.
The biological bacterial fertilizer for promoting the growth of the rhododendron delavayi tissue culture seedlings preferably comprises 10-15 parts by volume of a bacterial liquid of saccharomyces cerevisiae, more preferably 10-14 parts by volume of the bacterial liquid, and even more preferably 13 parts by volume of the bacterial liquid. In the invention, the viable count of the saccharomyces cerevisiae bacterial liquid is preferably more than or equal to 6 multiplied by 107CFU/mL; more preferably 7 to 9X 107CFU/mL; more preferably 8X 107CFU/mL. The saccharomyces cerevisiae can ferment and decompose carbohydrates around the root system of the rhododendron lapponicum, and decompose macromolecular carbohydrates into small molecules so as to be absorbed by the rhododendron lapponicum and promote the rhododendron lapponicum to grow strong seedlings. The invention has no special requirements on the type of the saccharomyces cerevisiae, and can be prepared by adopting the strains which are generally sold in the market in the field. In the embodiment of the invention, the saccharomyces cerevisiae is purchased from the institute of biotechnology, Beijing Beinana and Chuanglian union, and the strain number of the saccharomyces cerevisiae is BNCC 140110. In the invention, the culture temperature of the saccharomyces cerevisiae is preferably 27-30 ℃; further preferably 27-29 ℃; more preferably 28.5 ℃. In the invention, the culture time of the saccharomyces cerevisiae is preferably 24-36 h; further preferably 29-35 h; still more preferably 33 h. In the invention, the culture pH of the saccharomyces cerevisiae is preferably 4-6; further preferably 4.4 to 5.8; still more preferably 5.5. In the invention, the wine brewing is carried outThe yeast is preferably cultured on a YPD solid medium, the saccharomyces cerevisiae can be well cultured, and the saccharomyces cerevisiae is simple to prepare and low in cost. After the culture, the colony cultured by the solid culture medium is preferably washed by sterile water to obtain a bacterial liquid of the saccharomyces cerevisiae.
The biological bacterial fertilizer for promoting the growth of the tissue culture seedlings of rhododendron lapponicum comprises, by volume, preferably 5-10 parts of a bacterial liquid of trichoderma longibrachiatum, more preferably 6-9 parts, and even more preferably 8 parts. In the present invention, the viable count of the bacterial liquid of Trichoderma longibrachiatum is preferably not less than 4X 107CFU/mL; more preferably 5 to 8X 107CFU/mL; more preferably 6X 107CFU/mL. The trichoderma longibrachiatum has the effect of preventing and controlling pathogenic bacteria in the growth process of rhododendron lapponicum; the trichoderma longibrachiatum can generate active metabolites such as chitinase, glucanase, cellulase, protease and the like, and can damage the cell walls of rhododendron lapponicum pathogenic fungi; but also can secrete antibacterial protein or lyase to inhibit the infection of rhododendron lapponicum pathogenic bacteria; meanwhile, trichoderma longibrachiatum can also secrete extracellular enzymes such as glucosidase and the like to degrade antibiotic toxins generated by pathogenic bacteria and reduce the damage of rhododendron alpinum pathogenic bacteria to beneficial microorganisms of plant root systems. The invention has no special requirements on the species of the trichoderma longibrachiatum, and can be prepared by adopting the strains which are generally sold in the market in the field. In the embodiment of the invention, the trichoderma longibrachiatum is purchased from the institute of biotechnology, namely Beijing Beinana Chuanglian, and the strain number of the trichoderma longibrachiatum is BNCC 185642. In the invention, the culture temperature of the trichoderma longibrachiatum is preferably 26-29 ℃; further preferably 26.5-28.5 ℃; still more preferably 27 deg.c. In the invention, the culture time of the trichoderma longibrachiatum is preferably 60-72 h; further preferably 62-70 h; still more preferably 68 h. In the invention, the culture pH of the trichoderma longibrachiatum is preferably 4-6.5; further preferably 4.5 to 6; still more preferably 5.5. In the invention, the trichoderma longibrachiatum is preferably cultured on a PDA solid culture medium, trichoderma longibrachiatum can be well cultured, and the trichoderma longibrachiatum is simple and convenient to prepare and low in cost. After the culture, the present invention preferably uses a microorganism cultured on a solid mediumAnd (5) washing with bacterial water to obtain bacterial liquid of the trichoderma longibrachiatum.
The biological bacterial fertilizer for promoting the growth of the tissue culture seedlings of rhododendron lapponicum comprises, by volume, preferably 10-15 parts of bacterial liquid of paecilomyces lilacinus, more preferably 11-14 parts of bacterial liquid, and even more preferably 13 parts of bacterial liquid. In the present invention, the viable count of the bacterial liquid of the paecilomyces lilacinus is preferably not less than 6X 107CFU/mL; more preferably 6 to 8X 107CFU/mL; still more preferably 7X 107CFU/mL. The paecilomyces lilacinus belongs to endoparasitic fungi, has parasitism on pest eggs such as root-knot nematodes, cyst nematodes and the like in rhizosphere soil of rhododendron lapponicum, and has combined action with chitin and chitinase secreted by the parasite eggs so as to kill pests; meanwhile, the paecilomyces lilacinus can also generate chemical substances similar to indoleacetic acid to promote the growth and development of root systems, promotes the dissolution of insoluble phosphate through secreted organic acid, is convenient for the root systems of rhododendron lapponicum to absorb, and plays roles in preventing and controlling insect pests and strengthening seedlings. The invention has no special requirements on the type of the paecilomyces lilacinus and can adopt the strains which are generally sold in the market in the field. The paecilomyces lilacinus in the embodiment of the invention is purchased from Beijing Beinanna Chuanglian union biotechnology research institute, and the strain number of the paecilomyces lilacinus is BNCC 119092. In the invention, the culture temperature of the paecilomyces lilacinus is preferably 27-31 ℃; further preferably 27.5-30 ℃; still more preferably 29 deg.c. In the invention, the culture time of the paecilomyces lilacinus is preferably 36-50 h; further preferably 38-48 h; more preferably 45 h. In the invention, the culture pH of the paecilomyces lilacinus is preferably 4.8-6.2; further preferably 5 to 6; still more preferably 5.5. In the invention, the paecilomyces lilacinus is preferably cultured on a PDA solid culture medium, so that the paecilomyces lilacinus can be well cultured, and the paecilomyces lilacinus is simple to prepare and low in cost. After culturing, the bacterial colony cultured by the solid culture medium is preferably washed by sterile water to obtain the bacterial liquid of the paecilomyces lilacinus.
The biological bacterial fertilizer for promoting the growth of the rhododendron delavayi tissue culture seedlings preferably comprises 15-25 parts by volume of a bacterial liquid of bacillus polymyxa,more preferably 18 to 23 parts, and still more preferably 20 parts. In the present invention, the viable count of the bacterial liquid of Bacillus polymyxa is preferably not less than 5X 108CFU/mL; more preferably 6 to 9X 108CFU/mL; more preferably 8X 108CFU/mL. The bacillus polymyxa has the functions of biological nitrogen fixation, phosphorus dissolution and potassium dissolution, so as to promote the absorption of azalea on nitrogen, phosphorus and potassium in soil; meanwhile, the rhododendron mariae extract can secrete metabolites such as polymyxin and the like to play an antagonistic role on pathogenic microorganisms in soil and plants, and has a good inhibition effect on root rot and leaf spot of rhododendron lapponicum. The invention has no special requirements on the species of the bacillus polymyxa, and can adopt the strains which are generally sold in the market in the field. In the embodiment of the invention, the bacillus polymyxa is purchased from Beijing Beinana Chuanglian union biotechnology research institute, and the strain number of the bacillus polymyxa is BNCC 110073. In the invention, the culture temperature of the bacillus polymyxa is preferably 28-33 ℃; further preferably 28-32 ℃; more preferably 30 ℃. In the invention, the culture time of the bacillus polymyxa is preferably 30-58 h; further preferably 35-50 h; more preferably 40 h. In the invention, the rotation speed of the culture is preferably 150-190 r.min-1(ii) a More preferably 150 to 180 r.min-1(ii) a More preferably 160 r.min-1. In the invention, the pH value of the culture of the bacillus polymyxa is preferably 6.6-7.8; further preferably 7 to 7.8; still more preferably 7.2. In the invention, the inoculation amount of the bacillus polymyxa culture preferably accounts for 8-12% of the total volume of the culture medium; further preferably 9% to 11%; still more preferably 10%. In the invention, the bacillus polymyxa is preferably cultured on a beef extract peptone liquid culture medium, so that the bacillus polymyxa can be well cultured, and the bacillus polymyxa is simple to prepare and low in cost. The invention has no special requirement on the source of the beef extract peptone liquid culture medium, and can be realized by adopting a common commercially available culture medium.
The biological bacterial fertilizer for promoting the growth of the tissue culture seedlings of rhododendron lapponicum comprises 35-60 parts by volume of pumpkin juice, preferably 40-55 parts by volume of pumpkin juice and more preferably 50 parts by volume of pumpkin juice. In the present invention, the pumpkin juice is preferably prepared from pumpkin powder or fresh pumpkin. When the fresh pumpkin is used as the raw material to prepare the pumpkin juice, the preparation method of the pumpkin juice preferably comprises the following steps: crushing pumpkin pulp to obtain crushed pumpkin pulp; mixing the crushed pumpkin pulp with water, and steaming in water to obtain pumpkin juice. In the present invention, the mass of the crushed pumpkin pulp and the volume ratio of water are preferably 100 g: (300-400) mL, more preferably 100 g: 350 mL. In the invention, the cooking time is preferably 10-15 min, and more preferably 15 min. When the pumpkin juice is prepared by using the pumpkin powder as a raw material, the concentration of the pumpkin juice is preferably 250-330 g/L, more preferably 260-320 g/L, and even more preferably 280 g/L. The invention has no special requirement on the sources of the fresh pumpkins and the pumpkin powder, and can be prepared by adopting the conventional commercial products in the field. The pumpkin can provide early-stage nutrient substances for the rapid growth of microorganisms, shortens the recovery and growth time of the microorganisms when the biological bacterial manure is used, and has the characteristics of low cost, easy acquisition, environmental friendliness and the like.
According to the invention, through reasonable compounding of the microbial agent and screening of suitable nutrient substances, the biological bacterial manure capable of remarkably improving the survival rate of the transplanted tissue culture seedlings of alpine rhododendron and strengthening the seedlings is prepared, and high-quality development of the domestic alpine rhododendron industry is promoted.
The invention provides a method for promoting the growth of tissue culture seedlings of alpine rhododendrons, which applies the biological bacterial manure in the technical scheme to the tissue culture seedlings of alpine rhododendrons. The biological bacterial fertilizer can be mixed with other fertilizers for use, and can also be used independently. In the present invention, the biological bacterial manure is preferably revived before use. The resuscitation condition of the invention is preferably as follows: preheating the biological bacterial fertilizer in water at 10-20 ℃ for 60-90 min, and then transferring the biological bacterial fertilizer into water at 20-30 ℃ for placing for 120-180 min. In the invention, the preheating temperature is further preferably 12-18 ℃; more preferably still at 15 ℃. In the invention, the preheating time is further preferably 65-90 min; more preferably 80 min. After preheating, the method preferably transfers the waste water into water with the temperature of 20-30 ℃ and stands for 120-180 min; further preferably transferring the mixture into water with the temperature of 22-28 ℃ and placing the mixture for 120-160 min; further preferably, the mixture is transferred into water at 25 ℃ and placed for 150 min. In the invention, the biological bacterial fertilizer is preferably used before the rhododendron lapponicum is transplanted, and is applied for 1 time every 3-4 months after transplantation. The invention has no special requirement on the transplanting matrix and can adopt the conventional transplanting matrix in the field. In the invention, the application amount of the biological bacterial fertilizer is preferably 100-200 mL of the biological bacterial fertilizer per 5kg of the substrate; further preferably, 100-150 mL of biological bacterial fertilizer is applied to every 5kg of the substrate; even more preferably 150mL of biological bacterial manure is applied per 5kg of substrate. In the present invention, the amount of the top application is preferably the same as the amount applied before transplantation. The biological bacterial fertilizer is used before transplanting, so that beneficial bacteria in soil are increased, and the planting and survival of transplanted seedlings are met; the invention is applied for 1 time after transplanting, can meet the requirement of bacterial manure in the growth period of the seedling, is beneficial to strengthening the seedling and the root and improves the growth quality of the seedling.
For further illustration of the present invention, the following examples are provided to describe the biological bacterial manure and the method for promoting the growth of the tissue culture seedling of rhododendron lapponicum in detail, but they should not be construed as limiting the scope of the present invention.
Example 1
A biological bacterial fertilizer for promoting growth of tissue culture seedlings of rhododendron lapponicum comprises the following components in parts by volume: 11 parts of a bacterial liquid of saccharomyces cerevisiae, 8 parts of a bacterial liquid of trichoderma longibrachiatum, 11 parts of a bacterial liquid of paecilomyces lilacinus, 20 parts of a bacterial liquid of bacillus polymyxa and 50 parts of pumpkin juice;
the viable count of the saccharomyces cerevisiae bacterial liquid is 7 multiplied by 107CFU/mL; the viable count of the trichoderma longibrachiatum bacterial liquid is 5 multiplied by 107CFU/mL; the viable count of the paecilomyces lilacinus bacterial liquid is 6 multiplied by 107CFU/mL; the viable count of the bacillus polymyxa liquid is 6 multiplied by 108CFU/mL。
The preparation method comprises the following steps:
1) preparing sterile water, filling distilled water into a sterile wide-mouth bottle, placing in an autoclave, sterilizing at 120 deg.C for 10min, and cooling.
2) Cleaning raw materials: peeling the peel of the ripe pumpkin, removing seeds, grinding the rest pulp part into pumpkin juice by using a tissue refiner, adding 400mL of distilled water into every 100g of pumpkin pulp, boiling the ground pumpkin juice in boiling water at 100 ℃ for 15min in a water-separating way, cooling, and then putting into a plastic tube with a sterile cover for storage for later use.
3) Respectively carrying out recovery culture on saccharomyces cerevisiae, trichoderma longibrachiatum, paecilomyces lilacinus and bacillus polymyxa. The recovery culture condition of the saccharomyces cerevisiae adopts a YPD solid culture medium plate upper-scribing method for inoculation culture, the culture temperature is 28 ℃, the pH value is 5, and the culture time is 30 h; inoculating and culturing the trichoderma longibrachiatum by adopting a PDA solid medium plate scribing method, wherein the culture temperature is 27 ℃, the pH value is 6, and the culture time is 65 hours; the paecilomyces lilacinus is inoculated and cultured by a PDA solid medium plate scribing method, the culture temperature is 29 ℃, the pH value is 5.5, and the culture time is 40 h; the bacillus polymyxa is cultured by beef extract peptone liquid at the temperature of 30 ℃ and the rotating speed of 170 r.min and the pH value of 7-1The liquid loading amount is 80mL/250mL, the inoculation amount is 10% (volume fraction), and the culture time is 40 h. In this example, Saccharomyces cerevisiae, Trichoderma longibrachiatum, Paecilomyces lilacinus and Bacillus polymyxa were purchased from Beijing Ministry of Biotechnology, wherein the Saccharomyces cerevisiae strains are BNCC140110, Trichoderma longibrachiatum strain BNCC185642, Paecilomyces lilacinus strain BNCC119092 and Bacillus polymyxa strain BNCC 110073.
4) Washing the microorganism cultured in the solid culture medium in the third step with sterile water to obtain bacterial liquid, wherein the viable count of the bacterial liquid of Saccharomyces cerevisiae is 7 × 107CFU/mL, viable count of the bacterial liquid of Trichoderma longibrachiatum is 5 × 107CFU/mL, viable count of the bacterial liquid of the paecilomyces lilacinus is 6 multiplied by 107CFU/mL, viable count of Bacillus polymyxa bacterial liquid of 6X 108And (5) CFU/mL, respectively filling the bacterial liquid into a sterile wide-mouth plastic bottle for storage and standby.
5) Mixing the following components in parts by volume: each 100mL of the mixed bacterial liquid contains 11mL of saccharomyces cerevisiae, 8mL of trichoderma longibrachiatum, 11mL of paecilomyces lilacinus, 20mL of bacillus polymyxa and 50mL of pumpkin juice. And mixing to obtain the biological bacterial fertilizer for promoting the growth of the tissue culture seedlings of rhododendron lapponicum.
Example 2
A biological bacterial fertilizer for promoting growth of tissue culture seedlings of rhododendron lapponicum comprises the following components in parts by volume: 10 parts of a bacterial liquid of saccharomyces cerevisiae, 9 parts of a bacterial liquid of trichoderma longibrachiatum, 11 parts of a bacterial liquid of paecilomyces lilacinus, 22 parts of a bacterial liquid of bacillus polymyxa and 48 parts of pumpkin juice;
the viable count of the saccharomyces cerevisiae bacterial liquid is 7 multiplied by 107CFU/mL; the viable count of the trichoderma longibrachiatum bacterial liquid is 7 multiplied by 107CFU/mL; the viable count of the paecilomyces lilacinus bacterial liquid is 8 multiplied by 107CFU/mL; the viable count of the bacillus polymyxa liquid is 6 multiplied by 108CFU/mL。
The preparation method is the same as example 1.
Example 3
A biological bacterial fertilizer for promoting growth of tissue culture seedlings of alpine rhododendron comprises the following components in parts by volume: 14 parts of a bacterial liquid of saccharomyces cerevisiae, 7 parts of a bacterial liquid of trichoderma longibrachiatum, 12 parts of a bacterial liquid of paecilomyces lilacinus, 19 parts of a bacterial liquid of bacillus polymyxa and 48 parts of pumpkin juice;
the viable count of the saccharomyces cerevisiae bacterial liquid is 6 multiplied by 107CFU/mL; the viable count of the trichoderma longibrachiatum bacterial liquid is 5 multiplied by 107CFU/mL; the viable count of the paecilomyces lilacinus bacterial liquid is 7 multiplied by 107CFU/mL; the viable count of the bacillus polymyxa liquid is 6 multiplied by 108CFU/mL。
The preparation method is the same as example 1.
Comparative example 1
The biological bacterial fertilizer for promoting the growth of tissue culture seedlings of rhododendron lapponicum is the same as that in example 1, and is characterized in that: saccharomyces cerevisiae was not used.
The preparation method is the same as example 1.
Comparative example 2
The biological bacterial fertilizer for promoting the growth of tissue culture seedlings of rhododendron lapponicum is the same as that in example 1, and is characterized in that: paecilomyces lilacinus and Bacillus polymyxa were not used.
The preparation method is the same as example 1.
Comparative example 3
The biological bacterial fertilizer for promoting the growth of tissue culture seedlings of rhododendron lapponicum is the same as that in example 1, and is characterized in that: the pumpkin is replaced by the potato.
The preparation method is the same as example 1.
Comparative example 4
The biological bacterial fertilizer for promoting the growth of tissue culture seedlings of rhododendron lapponicum is the same as that in example 1, and is characterized in that: the pumpkin is replaced by the bean cake powder.
The preparation method is the same as example 1.
Comparative example 5
The biological bacterial fertilizer for promoting the growth of tissue culture seedlings of rhododendron lapponicum is the same as that in example 1, and is characterized in that: the pumpkin is replaced by the nitrogen-phosphorus-potassium compound fertilizer.
The preparation method is the same as example 1.
Example 4
In order to investigate the rationality of the microbial formula in the technical scheme, the microbial species in the formula are reduced, and the using effect of the bacterial manure in the formula after reduction is observed. Respectively selecting sterile tissue culture seedlings of Rhododendron delavayi, Rhododendron delavayi and Rhododendron delavayi as test objects, respectively setting 4 groups of test groups, respectively setting a control group, a biological bacterial fertilizer group, a bacteria reduction 1 group and a bacteria reduction 2 group, respectively setting 30 seedlings with the same growth vigor in each group, selecting humus soil under the Pleiodendron for a transplanting substrate, uniformly sterilizing at high temperature before use, and filling the sterilized seedlings into a flowerpot with the diameter of 20cm for later use. In the test, the control group uses the same amount of clear water to replace biological bacterial manure; the biological bacterial manure of the embodiment 1 of the invention is applied to a biological bacterial manure group, before use, 100mL of recovered biological bacterial manure is diluted by 2L of water, then is uniformly poured into 5kg of matrix, is uniformly stirred, and then seedlings are transplanted; the recovery method of the biological agent comprises the following steps: preheating the biological bacterial manure in water of 15 ℃ for 80min, and then transferring the biological bacterial manure into water of 25 ℃ for standing for 150 min; the group of bacteria reduction 1 is applied with the biological bacterial manure of the comparative example 1, and the recovery method and the application amount of the biological bacterial manure are the same as those of the biological bacterial manure group; and (3) applying the biological bacterial manure of the comparative example 2 to the group with the bacteria reduction 2, wherein the recovery method and the application amount of the biological bacterial manure are the same as those of the group with the biological bacterial manure. And other management and protection conditions are consistent, indexes of plant height, leaf area, total root length, dry matter content and chlorophyll content of leaves of the plants are measured 30 days after treatment of each group, the growth condition of seedlings is judged, and the detection results are shown in tables 1-4 and figures 1-3.
The details of the influence of different bacterial manure formula treatments on the growth conditions of the tissue culture seedlings of the rhododendron pulchrum are shown in figure 1; wherein the growth conditions of the rhododendron under the treatment of a control group, a bacteria-reducing group 2, a bacteria-reducing group 1 and a biological bacterial manure group for 30 days are sequentially arranged from left to right. The growth condition of the Rhododendron delavayi tissue culture seedling treated by the biological bacterial manure is shown in figure 2; wherein the growth conditions of the Rhododendron delavayi Franch under the treatment of the biological bacterial manure group and the control group of the invention are sequentially from left to right for 30 days. The growth condition of the rhododendron delavayi tissue culture seedling treated by the biological bacterial manure is shown in figure 3; wherein the growth conditions of rhododendron delavayi under the treatment of a control group and the biological bacterial manure group of the invention for 30 days are sequentially arranged from left to right.
TABLE 1 Effect of different bacterial manure formulations on the growth of tissue cultured seedlings of Rhododendron delavayi Franch
Figure BDA0003037104860000111
TABLE 2 influence of different bacterial manure formulations on growth of tissue-cultured seedlings of Rhododendron anthopogonoides
Figure BDA0003037104860000112
Figure BDA0003037104860000121
TABLE 3 influence of different bacterial manure formulations on the growth of tissue-cultured seedlings of Rhododendron delavayi Franch
Figure BDA0003037104860000122
TABLE 4 influence of different bacterial manure formulations on the growth of tissue-cultured seedlings of wide cup rhododendron
Figure BDA0003037104860000123
As can be seen from FIG. 1, the growth vigor of the biological bacterial manure treatment group is obviously superior to that of the control group, the bacteria reduction 2 group and the bacteria reduction 1 group, which indicates that the microbial species in the formula of the invention are all deficient. As can be seen from the results of fig. 2 and fig. 3, the growth vigor of the biological bacterial fertilizer treatment of the invention is significantly better than that of the control group, which indicates that the biological bacterial fertilizer of the invention has good promotion of the growth vigor of rhododendron lapponicum and good seedling strengthening effect. And by specifically combining tables 1 to 4, for different alpine rhododendron varieties, the effect of bacterial manure is obviously reduced by reducing the microbial varieties in the formula, and compared with a biological bacterial manure group, the bacterial manure group 1 and the alkaline bacteria group 2 have obvious differences in plant height, leaf area, total root length, dry matter content and chlorophyll SPAD value and are reduced along with the reduction of the microbial varieties, so that the defect of the microbial varieties in the formula is further illustrated.
Example 5
In order to investigate whether the pumpkin has substitutability, the pumpkin is substituted by the potato, the bean cake powder and the nitrogen-phosphorus-potassium compound fertilizer respectively. Taking Rhododendron delavayi, Rhododendron fasciatus, Rhododendron delavayi and Rhododendron delavayi sterile tissue culture seedlings as test objects, wherein each test is respectively provided with 4 groups of test groups, namely a control group, a biological bacterial fertilizer group, a potato group, a bean cake powder group and a nitrogen-phosphorus-potassium compound fertilizer group; and (3) selecting humus soil under the hollandia baileyi forest for transplanting substrates of 30 seedlings in each group, sterilizing at high temperature uniformly before use, and filling the seedlings into flowerpots with the apertures of 20cm for later use after sterilization. The control group in the experiment was replaced with an equal amount of water; the biological bacterial manure of the embodiment 1 of the invention is applied to a biological bacterial manure group, before use, 120mL of biological bacterial manure is revived and added into 2L of water, and the mixture is uniformly irrigated to 5kg of matrix and is uniformly mixed; the potato groups were applied with the formulation of comparative example 3; the legume meal group was applied with the formulation of comparative example 4; the nitrogen phosphorus potassium compound fertilizer group uses the formulation of the comparative example 5. Other management and protection conditions are consistent, and the content of alkaline hydrolysis nitrogen, quick-acting phosphorus, quick-acting potassium, the content of organic matters and the concentration of soluble ions in the soil in the matrix are measured at 10 days, 20 days and 30 days after treatment of each group; and (3) measuring the plant height, the leaf area, the total root length, the dry matter content and the chlorophyll content of the leaves of the plants 30 days after the treatment, judging the soil condition and the seedling growth condition, and obtaining the detection results shown in tables 5-9.
TABLE 5 Effect of different formulations on the physicochemical Properties of the culture substrate
Figure BDA0003037104860000131
Figure BDA0003037104860000141
As can be seen from the results in Table 5, the pumpkin is replaced by the potato powder, the bean cake powder and the nitrogen-phosphorus-potassium compound fertilizer in the substrate, and the comparison condition of the content of the alkaline hydrolysis nitrogen, the quick-acting phosphorus, the quick-acting potassium, the content of the organic matters and the concentration of the soluble ions in the treated substrate at 10 days, 20 days and 30 days is researched, and the result shows that the concentration of the soluble ions in the biological bacterial manure group is increased compared with that in the control group, but the increase amount does not exceed the tolerance maximum limit of the plants to the concentration of the soluble ions in the soil by 2.6ms cm-1Still being suitable for plant growth; meanwhile, the contents of organic matters, alkaline hydrolysis nitrogen, quick-acting phosphorus and quick-acting potassium are obviously higher than those of a control group, a potato group and a bean cake powder group, although the contents of the alkaline hydrolysis nitrogen, the quick-acting phosphorus and the quick-acting potassium are not greatly different at 10d and 20d compared with those of a nitrogen-phosphorus-potassium compound fertilizer group, the contents of the alkaline hydrolysis nitrogen, the quick-acting phosphorus and the quick-acting potassium are obviously higher than those of the nitrogen-phosphorus-potassium compound fertilizer group at 30d, and the formula of the invention has more lasting effect than that of a conventional inorganic fertilizer formula. The conclusion shows that the pumpkin has an irreplaceable effect in the formula.
TABLE 6 Effect of different formulations on the growth of tissue-cultured seedlings of Rhododendron delavayi Franch
Figure BDA0003037104860000142
7 influence of different formulas on growth of tissue culture seedlings of rhododendron pulchrum
Figure BDA0003037104860000143
Figure BDA0003037104860000151
TABLE 8 Effect of different formulations on Rhododendron delavayi tissue culture seedling growth
Figure BDA0003037104860000152
TABLE 9 Effect of different formulations on the growth of tissue-cultured seedlings of Rhododendron pulchrum
Figure BDA0003037104860000153
From the results in tables 6 to 9, it can be seen that the potato group, the bean cake powder group and the nitrogen-phosphorus-potassium compound fertilizer group are inferior to the biological bacterial fertilizer group in the four materials of Rhododendron delavayi, Rhododendron fasciatus, Rhododendron delavayi and Rhododendron delavayi. On the 5 indexes of plant height, leaf area, total root length, dry matter content and chlorophyll SPAD value, the biological bacterial manure group is obviously superior to a control group, a potato group, a bean cake powder group and a nitrogen-phosphorus-potassium compound fertilizer group, and the pumpkin in the formula disclosed by the invention is further proved to be non-replaceable.
Example 6
The influence of the biological bacterial fertilizer application, the conventional application and the organic application on the rhododendron lapponicum tissue culture seedling for promoting the growth of the rhododendron lapponicum tissue culture seedling provided by the invention is examined. Respectively selecting sterile tissue culture seedlings of Rhododendron delavayi, Rhododendron deworme, Rhododendron fasciatus, Rhododendron cupulatum and Rhododendron delavayi as test objects, respectively setting 6 groups of test groups, selecting humus soil under the Pleiodendron for a transplanting substrate, uniformly sterilizing at high temperature before use, and filling the sterilized seedlings into flowerpots with 20cm calibers for later use. In the test, the control group uses the same amount of clear water to replace biological bacterial manure; the biological bacterial manure of the embodiment 1 of the invention is applied to the biological bacterial manure group 1, 150mL of the biological bacterial manure is revived before use and then added into 2L of water, and the mixture is uniformly poured to 5kg of matrix and is uniformly stirred; biological bacterial manure 2 groups were applied with the biological bacterial manure of example 2 of the present invention; biological bacterial manure 3 groups were applied with the biological bacterial manure of example 3 of the present invention; the nitrogen-phosphorus-potassium compound fertilizer is applied by a conventional fertilizing group, and the proportion of nitrogen, phosphorus and potassium is 21: 7: 7, fertilizing amount of each plant is 200 g; the organic fertilization group applies fermented farmyard manure with the fertilizing amount of 200g per plant, and the farmyard manure is evenly mixed into a transplanting substrate before transplanting. Other management and protection conditions are consistent, indexes of plant height, leaf area, total root length, dry matter content and chlorophyll content of leaves are measured 30 days after treatment of each group, the growth condition of seedlings is judged, and detection results are shown in tables 10-14.
TABLE 10 Effect of different fertilizer types on Rhododendron delavayi tissue culture seedling growth
Figure BDA0003037104860000161
TABLE 11 Effect of different Fertilizer species on tissue culture seedling growth of Rhododendron dewdrop
Figure BDA0003037104860000162
Figure BDA0003037104860000171
TABLE 12 Effect of different Fertilizer classes on the growth of tissue-cultured seedlings of Azalea charantia
Figure BDA0003037104860000172
TABLE 13 Effect of different Fertilizer species on Rhododendron delavayi tissue culture seedling growth
Figure BDA0003037104860000173
TABLE 14 Effect of different Fertilizer classes on the growth of tissue-cultured seedlings of Rhododendron pulchrum
Figure BDA0003037104860000174
Figure BDA0003037104860000181
From the results in tables 10 to 14, it can be seen that for the Rhododendron delavayi Franch, Rhododendron deworme, Rhododendron odoratum, Rhododendron cupulatum and Rhododendron delavayi Franch, the seedling height, leaf area, total root length, dry matter content and SPAD value of the biological bacterial fertilizer group of the present invention are significantly superior to those of the control group, the conventional fertilizer group and the organic fertilizer group, and the biological bacterial fertilizer of the present invention significantly promotes the growth of tissue culture seedlings of Rhododendron delavayi Franch, Rhododendron deworme, Rhododendron odoratum, Rhododendron cupulatum and Rhododendron delavayi Franch.
Example 7
The influence of the biological bacterial fertilizer application, the conventional application and the organic application on the rhododendron lapponicum tissue culture seedlings when being popularized and used in a large area is investigated. Selecting a nonalongshan rhododendron aseptic tissue culture seedling as a test object, selecting a seedling culture base in a scientific research institute in a rhododendron management area in a test field, setting 4 test groups in a simple plastic greenhouse in a seedling transplanting growth environment, setting 5000 seedlings in each group with consistent growth vigor, selecting humus soil under a rhododendron of a transplanting substrate, sterilizing at high temperature uniformly before use, and filling the sterilized seedlings into flowerpots with the diameter of 20cm for later use. In the test, the control group uses the same amount of clear water to replace biological bacterial manure; the biological bacterial manure of the embodiment 1 of the invention is applied to the biological bacterial manure group 1, 150mL of the biological bacterial manure is revived before use and then added into 2L of water, and the mixture is uniformly poured to 5kg of matrix and is uniformly stirred; the nitrogen-phosphorus-potassium compound fertilizer is applied by a conventional fertilizing group, and the proportion of nitrogen, phosphorus and potassium is 21: 7: 7, fertilizing amount of each plant is 200 g; the organic fertilization group applies fermented farmyard manure with the fertilizing amount of 200g per plant, and the farmyard manure is evenly mixed into a transplanting substrate before transplanting. Other management and protection conditions are consistent, indexes of plant height, leaf area, total root length, dry matter content and chlorophyll content of leaves are measured 30 days after treatment of each group, the growth condition of seedlings is judged, and detection results are shown in table 15, and fig. 4 and 5.
TABLE 15 influence of different tissue culture seedling-hardening modes on the growth of tissue-cultured seedlings of Rhododendron delavayi Franch
Figure BDA0003037104860000182
From the results in table 15, it can be seen that, for the rhododendron nonalongatum, the seedling plant height, leaf area, total root length, dry matter content and SPAD value of the biological bacterial fertilizer group of the present invention are significantly better than those of the control group, the conventional fertilization group and the organic fertilization group in the area production practical use, and the biological bacterial fertilizer of the present invention significantly promotes the growth of the tissue culture seedling of the rhododendron nonalongatum. The growth condition of 30d after transplantation in large-area use is shown in figure 4; the development effect of the root system of the rhododendron nonalongatum treated by the biological bacterial manure is shown in figure 5. From the results of fig. 4 and 5, it can be known that the biological bacterial manure prepared by the invention in large area in actual production has significant growth promotion effect on the tissue culture seedlings of the rhododendron nonalongshan after transplantation, has significant effect on promoting the root development of the seedlings, significantly improves the number of the root systems and the growth quality of the root systems, and simultaneously has promotion effect on the overground part growth of the seedlings, significantly improves the number of the leaves, the area and the chlorophyll content, and improves the survival rate of the seedlings as a whole by nearly 1 time.
The results of the above embodiments show that the biological bacterial fertilizer for promoting the growth of the tissue culture seedlings of rhododendron lapponicum provided by the invention can significantly improve the survival rate and the strong seedlings of the transplanted tissue culture seedlings of rhododendron lapponicum, and promote the high-quality development of the rhododendron lapponicum industry.
The above embodiments are described in detail, but they are only a part of the embodiments of the present invention, rather than all embodiments, and other embodiments can be obtained without inventive step according to the embodiments, and the embodiments are within the scope of the present invention.

Claims (9)

1. The biological bacterial fertilizer for promoting the growth of tissue culture seedlings of rhododendron lapponicum is characterized by comprising the following components in parts by volume: 10-15 parts of a saccharomyces cerevisiae bacterial solution, 5-10 parts of a trichoderma longibrachiatum bacterial solution, 10-15 parts of a paecilomyces lilacinus bacterial solution, 15-25 parts of a bacillus polymyxa bacterial solution and 35-60 parts of pumpkin juice;
the viable count of the saccharomyces cerevisiae bacterial liquid is more than or equal to 6 multiplied by 107CFU/mL; the viable count of the trichoderma longibrachiatum bacterial liquid is more than or equal to 4 multiplied by 107CFU/mL; the viable count of the paecilomyces lilacinus liquid is more than or equal to 6 multiplied by 107CFU/mL; the viable count of the bacillus polymyxa liquid is more than or equal to 5 multiplied by 108CFU/mL。
2. The biological bacterial fertilizer according to claim 1, wherein the saccharomyces cerevisiae is cultured under the following conditions: the temperature is 27-30 ℃, the time is 24-36 h, and the pH is 4-6.
3. The biological bacterial fertilizer as claimed in claim 1, wherein the culture conditions of trichoderma longibrachiatum are as follows: the temperature is 26-29 ℃, the time is 60-72 hours, and the pH is 4-6.5.
4. The biological bacterial fertilizer according to claim 1, wherein the culture conditions of the paecilomyces lilacinus are as follows: the temperature is 27-31 ℃, the time is 36-50 h, and the pH is 4.8-6.2.
5. The biological bacterial fertilizer as claimed in claim 1, wherein the culture conditions of the bacillus polymyxa are as follows: the temperature is 28-33 ℃, the time is 30-58 h, and the pH is 6.6-7.8.
6. A method for promoting the growth of tissue culture seedlings of rhododendron lapponicum, which is characterized in that the biological bacterial fertilizer as defined in any one of claims 1 to 5 is applied to the tissue culture seedlings of rhododendron lapponicum.
7. The method of claim 6, wherein the biological bacterial manure is used in a mixture with other fertilizers or used alone.
8. The method according to claim 6, wherein the biological bacterial manure is used before the rhododendron lapponicum is transplanted and is applied for 1 time after the rhododendron lapponicum is transplanted every 3-4 months; the application amount of the biological bacterial fertilizer is 100-200 mL of the biological bacterial fertilizer applied to every 5kg of the substrate.
9. The method of claim 6, wherein the biological bacterial manure is resuscitated prior to use; the resuscitation conditions are as follows: preheating the biological bacterial fertilizer in water at 10-20 ℃ for 60-90 min, and then transferring the biological bacterial fertilizer into water at 20-30 ℃ for placing for 120-180 min.
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