CN103820348B - The production method of one strain plant growth-promoting bacteria and microbial inoculum thereof and application - Google Patents
The production method of one strain plant growth-promoting bacteria and microbial inoculum thereof and application Download PDFInfo
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- CN103820348B CN103820348B CN201210461410.7A CN201210461410A CN103820348B CN 103820348 B CN103820348 B CN 103820348B CN 201210461410 A CN201210461410 A CN 201210461410A CN 103820348 B CN103820348 B CN 103820348B
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Abstract
The present invention relates to the bacillus amyloliquefaciens of a strain controlling plant diseases, Promoting plant growth, and the Low-cost production method of microbial inoculum, belong to microbial pesticide and microbial fertilizer technical field.Does is the bacterial strain that the present invention adopts separation screening plant height effect plant growth-promoting bacteria---a bacillus amyloliquefaciens (Bacillus out from pasture and water rhizosphere soil? amyloliquefaciens) A3 bacterial strain, this bacterial strain is deposited in China Committee for Culture Collection of Microorganisms's common micro-organisms center, does is deposit number: CGMCC? No.6676.The present invention utilizes Sweet Potato Starch Processing Waste Water as fermention medium, large scale culturing bacillus amyloliquefaciens, preparation has the microbial inoculum of controlling plant diseases, Promoting plant growth, this explained hereafter is with low cost, simple to operate, Sweet Potato Starch Processing Waste Water can also be made to get utilization, and decreasing pollution discharges.The microbial inoculum that the present invention produces is used for the production of tealeaves and vegetables, not only can improve output, and can improve the quality of tea place and vegetables, demonstrate wide application prospect.
Description
Technical field:
The present invention relates to the bacillus amyloliquefaciens of a plant height effect antagonistic phytopathogen, Promoting plant growth, and Low-cost production method and using method, belong to microbial pesticide and microbial fertilizer technical field.
Background technology:
Bacillus amyloliquefaciens (Bacillusamyloliquefaciens) possesses the diastatic potentiality of product with it and gains the name.Bacillus amyloliquefaciens is widely distributed at occurring in nature, is a kind of important biocontrol microorganisms, and it suppresses plant pathogenetic bacteria, fungi, virus, nematode by producing microbiotic, antibacterial protein or polypeptide isoreactivity material, thus plays the effect of biological control; Simultaneously it is also the important plant rhizosphere growth-promoting bacterium of a class, shows the effect to various plants growth promoting effects.Such as, it is active that Idriss etc. report that bacillus amyloliquefaciens FZB45 bacterial strain has phytase (phytase), can grow (Microbiology, 2002,148:2097-2109) by stimulating plant under the condition of scarce phosphorus.Shen Lili etc. report that bacillus amyloliquefaciens Ba33 bacterial strain can promote tobacco leaf production and suppress the propagation (Jilin Auto Industry, 2010,32:383 ~ 386) of tobacco mosaic virus (TMV).Lee is superfine there was reported the Bacillus amyloliquefaciens strain DC1(ACTA Scientiae Circumstantiae that a strain has algae-lysing, 31:1602-1608).
Chinese invention patent (application number: disclose a bacillus amyloliquefaciens Lx-11 bacterial strain 201010518034.1), deposit number is CGMCCNo.3789, good prevention effect is had to bacterial leaf streak of rice, the substratum cultivating the employing of this bacterial strain is: soyflour 1%, soybean cake powder 2%, starch 0.5%, yeast extract paste 0.2%, corn steep liquor 0.22%, sodium-chlor 0.1%, Calcium hydrogen carbonate 0.18%.Chinese invention patent (application number: disclose a bacillus amyloliquefaciens YN-1 bacterial strain 200810231406.5), deposit number is CGMCCNo.2328, can effectively prevent and treat the fungal disease such as apple marssonina leaf spot, wheat hypochnus, and filtered out industrial substratum with low cost, composition comprises: glucose 8 ~ 12g/L, Semen Maydis powder 10 ~ 16g/L, soybean cake powder 10 ~ 16g/L.Chinese invention patent (application number: disclose a bacillus amyloliquefaciens HN bacterial strain 200910070709.8), deposit number is CGMCCNo.3261, the nitric nitrogen in aquaculture water, nitrite nitrogen, COD equal size can be made obviously to reduce, and the cultivation of this bacterial strain adopts broth culture.Chinese invention patent (application number: disclose a plants endogenetic bacillus amyloliquefaciens TF28 bacterial strain 201010264520.5), deposit number is CGMCCNo.4038, it can in plant Colonization inside plants, breeding and conduction, antibacterial protein and lipopeptide antibiotic two kinds of antimicrobial substances can be produced, have disease prevention growth-promoting effect, the every 1000mL of its liquid nutrient medium soaks powder by 10g Tryptones, 5g yeast, 10gNaCl is formulated.Chinese invention patent (application number: disclose a bacillus amyloliquefaciens EA19 bacterial strain 200910273498.8), deposit number is CCTCCM209316, to Cochliobolus sativus, Root Rot of Wheat leaf spoting bacteria, gaeumannomyces graminis, wheat sclerotium rolfsii and rhizoctonia cerealis, all there is obvious bacteriostatic activity, its fermention medium forms: glucose 20 grams, extractum carnis 8 grams, L-sodium 5 grams, MgSO
40.5 gram, KCl0.5 gram, KH
2pO
41 gram, FeSO
46H
2o0.15 milligram, MnSO
4h
2o5 milligram, CuSO
45H
2o0.16 milligram, 1000 milliliters, water.Chinese invention patent (application number: disclose a bacillus amyloliquefaciens B-004 bacterial strain 201110095116.4), deposit number is CGMCCNo.4557, it is adopted rear early epidemic germ to tomato and has obvious antagonistic effect, in its substratum, carbon source can be Zulkovsky starch, lotus root starch, glucose, lactose, trehalose, sucrose etc., and nitrogenous source can be peptone, extractum carnis, yeast extract paste, NH
4nO
3, NH
4cl, (NH
4)
2sO
4deng, other nutrition sources can be the metal-salt etc. containing potassium, iron, zinc, sodium, manganese, magnesium and calcium and so on.
In sum, the bacterial strain with the bacillus amyloliquefaciens of antagonism Plant diseases is more, and its cultural method is also different, but, have no the bacterial strain for Tea Production, also have no the report adopting Sweet Potato Starch Processing Waste Water to cultivate bacillus amyloliquefaciens.
Summary of the invention:
The object of this invention is to provide the bacillus amyloliquefaciens of a plant height effect antagonistic phytopathogen, Promoting plant growth, and provide a kind of and can produce the method for this microbiobacterial agent and the using method of microbial inoculum by recycling Sweet Potato Starch Processing Waste Water.
Bacterial strain used in the present invention imitates bacillus amyloliquefaciens (Bacillusamyloliquefaciens) the A3 bacterial strain of Promoting plant growth, antagonistic phytopathogen by contriver's separation screening plant height out from pasture and water rhizosphere soil, this bacterial strain is deposited in China Committee for Culture Collection of Microorganisms's common micro-organisms center on October 15th, 2012, deposit number is: CGMCCNo.6676, depositary institution address: Datun Road, Chaoyang District, Beijing City, Institute of Microorganism, Academia Sinica, postcode: 100101.
The production method of bacillus amyloliquefaciens microbiobacterial agent of the present invention is as follows:
1, Sweet Potato Starch Processing Waste Water pre-treatment: regulate the pH of waste water to 6.8 ~ 7.7, in 110 ~ 126 DEG C of sterilizings 10 ~ 35 minutes with aqueous sodium hydroxide solution or liming.
2, liquid seeds preparation: the bacillus amyloliquefaciens inoculating slant preservation in the triangular flask that the good Sweet Potato Starch Processing Waste Water of pre-treatment is housed, in 25 ~ 37 DEG C, 120 ~ 200 revs/min of shaking culture 24 ~ 36 hours, obtain liquid seeds.
3, liquid state fermentation: inoculate the liquid seeds that volume ratio is 2% ~ 15% to being equipped with in the good Sweet Potato Starch Processing Waste Water fermentor tank of pre-treatment, in 25 ~ 37 DEG C, logical sterile air is cultivated 15 ~ 36 hours; Cultured fermentation liquor count plate, viable count reaches 1.5 × 10
9more than CFU/ milliliter, gained fermented liquid directly can use as liquid bacterial agent.
4, solid fungicide preparation: to cross biological carbon (the remaining solid matter after biomass pyrolytic of 30 mesh sieves, main component is carbon) be carrier, according to fermented liquid: biological carbon is the weight ratio of 1: 1 ~ 1: 5, fermented liquid is added in biological carbon, fully mix, obtain solid-state microbial inoculum, air seasoning is to microbial inoculum water content less than 30%, packaging, leaves shady and cool dry place in, for subsequent use.
The using method of bacillus amyloliquefaciens microbiobacterial agent of the present invention is as follows:
1, fermented liquid or solid fungicide irrigation water being diluted to viable count is 5 × 10
6cFU/ milliliter ~ 5 × 10
7cFU/ milliliter, mixes, and applies in the soil near crop root according to the consumption ditch of every mu of 1 ~ 3 cubic metre of diluent;
2, fermented liquid or solid fungicide are added in fertilizer, mix, make the bacillus amyloliquefaciens viable count in fertilizer be 5 × 10
6cFU/ gram ~ 1 × 10
8cFU/ gram, in being manured into soil as base manure according to the consumption of 1 ~ 3 ton every mu.
Embodiment:
Can promote that crop is to the absorption of nutrient and utilization by bacillus amyloliquefaciens provided by the invention, and effectively can prevent and treat multiple fungi plant disease, the microbiobacterial agent that technical scheme provided by the invention is produced is applied to the usage quantity that agriculture production can reduce chemical fertilizer and agricultural chemicals, thus improves farm environment quality; And production cost is very low, high-concentration waste water is got utilization, decreasing pollution thing discharges.Embodiment is as follows:
Embodiment 1: Sweet Potato Starch Processing Waste Water produces bacillus amyloliquefaciens microbiobacterial agent
Sweet Potato Starch Processing Waste Water pre-treatment: regulate wastewater pH to 7.5, in 115 DEG C of sterilizings 20 minutes with liming.
Prepared by liquid seeds: the bacillus amyloliquefaciens inoculating slant preservation in the triangular flask that the good Sweet Potato Starch Processing Waste Water of pre-treatment is housed, and in 32 DEG C, 150 revs/min of shaking culture 26 hours, obtain liquid seeds.
Liquid state fermentation: inoculate 5%(volume ratio to being equipped with in the good Sweet Potato Starch Processing Waste Water fermentor tank of pre-treatment) liquid seeds, in 32 DEG C, logical sterile air cultivates 20 hours; Cultured fermentation liquor count plate, viable count reaches 5.5 × 10
9cFU/ milliliter.
Prepared by microbial inoculum: to cross the biological carbon of 30 mesh sieves for carrier, according to fermented liquid: biological carbon is the weight ratio of 1: 2, is added to by fermented liquid in biological carbon, fully mix, obtain solid-state microbial inoculum, air seasoning is to microbial inoculum water content less than 30%, packaging, leaves shady and cool dry place in.
Embodiment 2: bacillus amyloliquefaciens microbiobacterial agent is applied to organic tea and produces
Select uniform experimental field about 2 mu of tea tree growing way at organic tea garden, experiment tea tree and other tea trees of base manage and keep always, just apply fertilizer different.Experimental field be divided into 5 pieces, corresponding 5 process respectively, have the isolation strip being provided with about 2 meters between each process, by microbial inoculum with tap water dilution or directly by fermented liquid dilution use, 5 process are respectively clear water contrast (CK), starch wastewater contrast (FCK), high density (2.5 × 10
7cFU/ milliliter) ditch spread (H), middle concentration (1.8 × 10
7cFU/ milliliter) ditch spread (M), lower concentration (1.2 × 10
7cFU/ milliliter) ditch spread (L), use according to the amount of the microbial inoculum diluents of 3 liters every square metre.Each process uses twice microbial inoculum in an identical manner in the winter time, to gather spring tealeaves, result shows: the contrast spring tea output using wastewater dilution liquid contrasts without significant difference with clear water, and use the spring tea output of microbial inoculum significantly to increase, wherein, the spring tea output using middle concentration and high density microbial inoculum is suitable, all than clear water contrast volume increase more than 65%, uses the spring tea output of lower concentration microbial inoculum also than clear water contrast volume increase about 30%.And aminoacids content is significantly higher than clear water contrast in the tealeaves using microbial inoculum.
Embodiment 3: bacillus amyloliquefaciens microbiobacterial agent is used for preventing cucumber samping off
Cucumber samping off is the disease caused by fungal infection.Gather the soil of planting for many years in the warmhouse booth of cucumber, mix, load in flowerpot, 2 groups of flowerpots are set, often organize 10 basins, as in the greenhouses of 15 DEG C ~ 20 DEG C, keep higher levels of humidity and lower intensity of illumination.The 1st group of direct sowing cucumber seed of flowerpot, 15, every basin, in contrast.Admix the bacillus amyloliquefaciens microbiobacterial agent that weight ratio is 0.2% in 2nd multicast kind cucumber seeds forward direction soil, then 15 seeds sowed by every basin.Statistics sum after coming up, after seedling grows 1 true leaf, statistics lodging seedling sum before 2 true leaves.Result shows, contrast seedling lodging rate more than 40%, and adds the experimental group of microbial inoculum, and seedling lodging rate is lower than 5%.
Claims (3)
1. a strain have controlling plant diseases and Promoting plant growth bacillus amyloliquefaciens (
bacillusamyloliquefaciens), it is characterized in that: this bacillus amyloliquefaciens is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center, deposit number: CGMCCNo.6676.
2. adopt the bacillus amyloliquefaciens described in claim 1 to produce the method for microbiobacterial agent, comprise the steps:
(1) Sweet Potato Starch Processing Waste Water pre-treatment: regulate the pH of waste water to 6.8 ~ 7.7, in 110 ~ 126 DEG C of sterilizings 10 ~ 35 minutes with aqueous sodium hydroxide solution or liming;
(2) liquid seeds preparation: the bacillus amyloliquefaciens inoculating slant preservation in the triangular flask that the good Sweet Potato Starch Processing Waste Water of pre-treatment is housed, in 25 ~ 37 DEG C, 120 ~ 200 revs/min of shaking culture 24 ~ 36 hours, obtain liquid seeds;
(3) liquid state fermentation: inoculate the liquid seeds that volume ratio is 2% ~ 15% to being equipped with in the good Sweet Potato Starch Processing Waste Water fermentor tank of pre-treatment, in 25 ~ 37 DEG C, logical sterile air is cultivated 15 ~ 36 hours; Cultured fermentation liquor count plate, viable count reaches 1.5 × 10
9more than CFU/ milliliter, gained fermented liquid directly can use as liquid bacterial agent;
(4) solid fungicide preparation: to cross biological carbon (the remaining solid matter after biomass pyrolytic of 30 mesh sieves, main component is carbon) be carrier, according to fermented liquid: biological carbon is the weight ratio of 1: 1 ~ 1: 5, fermented liquid is added in biological carbon, fully mix, obtain solid-state microbial inoculum, air seasoning is to microbial inoculum water content less than 30%, packaging, leaves shady and cool dry place in.
3. the using method of the microbiobacterial agent produced by bacillus amyloliquefaciens described in claim 2, comprising:
(1) fermented liquid or solid fungicide irrigation water being diluted to viable count is 5 × 10
6cFU/ milliliter ~ 5 × 10
7cFU/ milliliter, mixes, and applies in the soil near crop root according to the consumption ditch of every mu of 1 ~ 3 cubic metre of diluent; Or
(2) fermented liquid or solid fungicide are added in fertilizer, mix, make the bacillus amyloliquefaciens viable count in fertilizer be 5 × 10
6cFU/ gram ~ 1 × 10
8cFU/ gram, in being manured into soil as base manure according to the consumption of 1 ~ 3 ton every mu.
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| CN110257366B (en) * | 2019-07-05 | 2021-08-20 | 中国农业科学院烟草研究所(中国烟草总公司青州烟草研究所) | Method for preventing and treating tobacco diseases by using bacillus subtilis based on biochar |
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