CN101942404B - Bacillus cereus and application thereof in promoting growth of laxus malrel - Google Patents
Bacillus cereus and application thereof in promoting growth of laxus malrel Download PDFInfo
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- CN101942404B CN101942404B CN2010102212181A CN201010221218A CN101942404B CN 101942404 B CN101942404 B CN 101942404B CN 2010102212181 A CN2010102212181 A CN 2010102212181A CN 201010221218 A CN201010221218 A CN 201010221218A CN 101942404 B CN101942404 B CN 101942404B
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Abstract
The invention discloses a bacillus cereus CLY07 which is separated from laxus malrel rhizosphere and can degrade lecithin. The classification designation is bacillus cereus; and the bacillus cereus is collected in China Center for Type Culture Collection (CCTCC), the collection number is CCTCC NO: M 2010157, and the collection date is June 24th, 2010. The invention also discloses an application of the bacillus cereus in promoting growth of laxus malrel. The CLY07 strain has high capacity for degrading lecithin; when the CLY07 microbial inoculum is used for inoculating the seedling plants of laxus malrel, the result indicates that the microbial inoculum can obviously promote the growth of laxus malrel seedlings; and therefore, the invention provides excellent strain resources for developing phosphobacteria fertilizer for laxus malrel.
Description
Technical field
The invention belongs to the microbial fertilizer technical field in the biological fertilizer field, be specifically related to a kind of bacillus cereus and the application in promoting the southerm yew growth thereof.
Background technology
Southerm yew (Taxus chinensis var.mairei) is the taxaceae Chinese yew genus plants; Be the species the widest, that growth is the fastest that distribute in this genus; Be widely distributed in the mountain region (Yu Wanjun etc., 1978) of China Zhejiang, Taiwan, Fujian, Jiangxi, Guangdong North, the north, Guangxi and northeast, Henan, Hubei and ground 600m～1200m such as western part, Hunan, SOUTH OF GANSU, Sichuan, Guizhou and northeast of Yunnan.Its tree crown is graceful, and material is hard, water is wet is stale-proof, be integrate view and admire, material is with, medicinal rare tree.
Chinese yew genus plants is because of extracting extremely common people's the concern of anti-cancer herb activeconstituents taxol.Taxol is one of at present best in the world cancer therapy drug.The present stage production of taxol utilizes the Taxus wildwood to supply raw materials mostly, but it is lower to belong to seeds natural propagation updating ability, and wild resource is very rare, can not satisfy the demand on the market far away.Because long-term serious felling causes southerm yew wildwood quantity to reduce sharply, Critical Condition (Pan's sign, 2004 have been at present; Wang Changteng, 2005).Through building large-area southerm yew artificial forest, can effectively protect this rare tree and improve the ecological environment, satisfy the required Sustainable development with the taxol industry of existing market.But because this platymiscium is tighter to environmental requirement, conventional artificial cultivation surviving rate is lower, and poor growth can not satisfy the demand in market far away.(2007) such as Jin nation's celebratings discover that using chemical fertilizer can significantly promote the artificial southerm yew growth of young plantations of building.Yet along with increasing sharply of global energy crisis and chemical fertilizer usage quantity, its negative impact is obvious day by day.Therefore, seek new fertilizer, especially biomass fertilizers receives much attention with the research that substitutes or partly substitute chemical fertilizer.
Phosphorus is being brought into play important effect in plant growth and development process, be one of essential important nutritive element of plant.The most of soil available phosphorus content of China is very low, and soil phosphorus mainly exists with the organic and inorganic phosphide state that plant can not absorb.Wherein, Organophosphorous compounds are important component parts of soil phosphorus, account for 1/3～1/2 of soil total phosphorus content.Organophosphorus in the soil can not directly be absorbed by plant, and they are progressively degraded under the effect of the enzyme of microorganism secretion, and the phosphorus that finally discharges utilizable inorganic states just can be by the plant absorbing utilization, and this process also is called as the mineralization of organophosphorus.Can the phosphorus that plant is difficult to absorb be converted into the mikrobe that can absorb form and be called phosphate solubilizing bacteria or phosphorus-solubilizing bacteria, mikrobe that can the mineralising organo phosphorous compounds is referred to as to separate the organophosphorus mikrobe.Research shows that soil is separated the organophosphorus mikrobe and generally accounted for 30%～60% of soil microorganisms sum.Compare with fungi, actinomycetes, the plant rhizosphere bacterium is considered to the most effectively separate the organophosphorus mikrobe.The former Russian scholar Meng Jinna (1935) is separated to a strain very capable bacillus megaterium (the Bacillus megatherium var.phosphaticum) bacterial strain of separating with Yelkin TTS that reduces nucleic acid first from soil, make P behind the inoculation soil
2O
5Content has improved more than 15%, this bacterial strain and in nineteen forty-seven mass production use.Nineteen fifty, China preceding northeast institute of agricultural sciences isolates the bacillus megaterium that can decompose organophosphorus from the black earth of northeast.After the eighties in 20th century, domestic many units have developed the compound phosphate solubilizing bacteria preparation of being made up of multiple genus bacillus in succession, and the research and the exploitation of microbial fertilizer have obtained great attention.The degrading organic phosphor bacterium of report mainly contains bacillus (Bacillus) both at home and abroad at present; Pseudomonas (Pseudomonas); Escherichia (Escherichia); Erwinia (Erwinia); Agrobacterium (Agrobacterium); Serratia (Serratia); Flavobacterium (Flavobacterium); Enterobacteria belongs to (Enterbacter); Micrococcus sp (Micrococcus); Azotobacter (Azotobacter); Salmonella (Salmonella); Chromobacterium (Chromobacterium); Alcaligenes (Alcaligenes); Arthrobacter (Arthrobacter); Sulfur oxidizing bacterium belongs to (Thiobacillusthiooxidans) and many Thiobacillus tens genus such as (Thiobacillu).
Can find out, the efficient phosphate solubilizing microorganism of screening from soil, development phosphorus decomposing microbial inoculum is manured into soil, and insoluble phosphorus is the low Sustainable development approach that drops into high production for the plant absorbing utilization in the activating soil.But discover that mikrobe has the intensive rhizosphere effect, under different soils, Different Crop rhizosphere and the varying environment condition, the kind and the quantity of separating the organophosphorus bacterium are all different.Therefore; Good in order to obtain with the southerm yew affinity; What be easy to rhizosphere colonization efficiently separates the organophosphorus bacterium; The present invention has separated plant height effect and has separated organophosphorus bacillus cereus (Bacillus cereus) CLY07 from China's southerm yew rhizosphere soil, show that through inoculation test it has obvious growth-promoting functions to southerm yew.At present, do not see that both at home and abroad southerm yew separates the report of organophosphorus bacterium aspect.
Summary of the invention
Technical problem to be solved by this invention provides a kind of bacillus cereus of southerm yew rhizosphere efficiently degrading organophosphorus thing.
The technical problem that the present invention also will solve provides the application of above-mentioned bacillus cereus in promoting the southerm yew growth.
In order to solve the problems of the technologies described above, the technical scheme that the present invention adopts is following:
From Shanxi Province's Changzhi City forestry bureau southerm yew rhizosphere soil; Screening has obtained bacillus cereus (Bacillus cereus) CLY07 of a high-efficiency degradation Yelkin TTS; Be preserved in Chinese typical culture collection center C CTCC, address: China. Wuhan. Wuhan University, postcode 430072; Deposit number: CCTCC NO:M 2010157, preservation date: on June 24th, 2010.
The main biological property of CCTCC NO:M 2010157 bacterial strains: on the beef-protein medium flat board, bacterium colony is bigger, oyster white, circle, tiling, neat in edge, bacterium colony surface drying, tarnish; Thalline is shaft-like, blunt garden, two ends, Gram-positive, about 0.95 * 1.45 μ m of size～2.50 * 4.10 μ m, central spore, peritrichous; Aerobic, oxydase reaction is negative, and catalase test is positive; The test of 3% Pottasium Hydroxide is negative, and the starch hydrolysis experiment is negative, ability glucose fermentation, D-ribose, SANMALT-S, pectinose, wood sugar, sucrose, trehalose, fructose; And produce acid, and unfermentable lactose, cottonseed sugar, rhamnosyl, D-semi-lactosi, melizitose, melibiose, Vitamin C2, cellobiose, N.F,USP MANNITOL, gelatin hydrolysis is negative; The nitrate reductase test is positive; Methyl red, VP feminine gender, indoles is positive, and the Citrate trianion growth test is negative; Through parasporal crystal dyeing, do not see that parasporal crystal occurs.
CCTCC NO:M 2010157 bacterial strain 16SrDNA gene orders are seen shown in the SEQ ID No.1.
Sequence in survey 16SrDNA sequence and the GenBank DB is carried out the BLAST comparison.The result shows that CLY07 bacterial strain and genus bacillus (Bacillus) homology is all very high, and wherein the similarity with Bacillus cereus is 100%.Combining form, physiological and biochemical property and 16SrDNA sequential analysis are accredited as bacillus cereus (Bacillus cereus).
The potted plant inoculation test of CCTCC NO:M 2010157 bacterial strain microbial inoculums shows that this microbial inoculum has obviously promoted growing of southerm yew.
Beneficial effect: bacillus cereus of the present invention can produce obviously molten phosphorus circle in periphery of bacterial colonies under the egg plate culture condition; Bacillus cereus of the present invention has better degradation effect to Yelkin TTS under the liquid oscilaltion culture condition, compare significant difference with contrast (CK); Bacillus cereus of the present invention is processed microbial inoculum inoculation southerm yew seedling, and the result shows that this microbial inoculum obviously promotes growing of southerm yew seedling.Therefore, the present invention provides good strain resource for exploitation southerm yew phosphor bacterium fertilizer.
Description of drawings
Bacillus cereus (Bacillus cereus) CLY07; Be preserved in Chinese typical culture collection center C CTCC, address: China. Wuhan. Wuhan University, postcode 430072; Deposit number: CCTCC NO:M 2010157, preservation date: on June 24th, 2010.
Fig. 1 is that the CLY07 bacterial strain dissolves the phosphorus circle on egg plate.
Fig. 2 is that the CLY07 bacterial strain is to the Yelkin TTS degradation capability.
Embodiment
According to following embodiment, can make those skilled in the art understand the present invention better.Embodiment is described only to be used to explain the present invention, and the present invention that should also can not limit in claims to be described in detail.
The dull and stereotyped phosphorus decomposing ability of embodiment 1:CLY07 is measured.
Egg yolk medium: Carnis Bovis seu Bubali cream 3.0g, peptone 10.0g, sodium-chlor 5.0g; Zero(ppm) water 1000mL, agar 15.0～18.0g, pH 7.0; Sterilization when treating that temperature is reduced to 50 ℃ of left and right sides, adds 3～4mL fresh-laid egg yellow liquor (yolk mixes with the SPSS equal-volume) among every 50mL; Mixing falls dull and stereotyped.
The CLY07 bacterial strain of activation 2 times is connected on the egg plate with sterilization toothpick point, cultivates after 3 days for 30 ℃, measure and dissolve phosphorus circle and colony diameter, and calculate the ratio (Fig. 1) that dissolves phosphorus circle and colony diameter.
Find out that from Fig. 1 the CLY07 bacterial strain can produce obviously opaque molten phosphorus circle after cultivating on the egg plate.Through measuring and calculating, dissolving the phosphorus loop diameter is 27.81mm, and colony diameter is 5.02mm, dissolves phosphorus circle and bacterium colony than up to 5.54.
Embodiment 2:CLY07 liquid phosphorus decomposing ability is measured.
The Meng Jinna substratum: glucose 10.0g, ammonium sulfate 0.5g, sodium-chlor 0.3g, Repone K 0.3g, MAGNESIUM SULPHATE HEPTAHYDRATE 99.5 0.3g, iron vitriol 0.03g, four water manganous sulfate 0.03g, Yelkin TTS 0.2g, lime carbonate 5.0g, zero(ppm) water 1000mL, pH 7.0
CLY07 inoculation NA substratum (Carnis Bovis seu Bubali cream 3.0g, peptone 10.0g, sodium-chlor 5.0g, zero(ppm) water 1000mL with activation 2 times; Agar 18.0g, pH 7.2～7.4) in, cultivate after 2 days for 30 ℃; The aseptic water washing thalline is processed bacteria suspension, gets the 1mL bacteria suspension in the 100mL triangular flask that contains 50mL Meng Jinna substratum, is contrast (CK) with the Meng Jinna substratum that connects 1mL sterilization bacteria suspension; Each handles 3 repetitions, and 30 ℃, behind the 200r/min shaking culture 72h; 4 ℃ of fermented liquids, the centrifugal 10min of 10000r/min, molybdenum antimony resistance colorimetric method is measured titanium pigment content (Fig. 2) in the fermented liquid.
Can find out that from Fig. 2 titanium pigment content is 2.05mgL in the inoculation CLY07 bacterial strain fermentation liquor
-1, be contrast (0.31mgL
-1) 6.61 times.Show that to sum up the CLY07 bacterial strain has stronger degradation capability to Yelkin TTS.
The test of embodiment 3:CLY07 greenhouse pot culture.
After CLY07 activation 2～3 times, be inoculated in a small amount of thalline of transfering loop picking and contain in 50mL NB substratum (sodium-chlor 5g, zero(ppm) water 1000mL, pH 7.2～7.4 for Carnis Bovis seu Bubali cream 3g, peptone 10g) the 100mL triangular flask, 30 ℃, 200r/min shaking culture 48h.Fermented liquid (4 ℃, 6000r/min) centrifugal 5min, behind SPSS rinse thalline 3 times, SPSS is regulated bacteria suspension (10
8Cfu/mL) process microbial inoculum.Inoculation southerm yew (seedling age 60 days) is contrast with the equivalent SPSS, and inoculum size is the 5mL/ strain.8 repetitions of every processing place the greenhouse unified management, and illumination is 12h/ days, waters in good time.
150 days growth situation (seeing table 1) of southerm yew seedling.Can find out that from table 1 inoculation CLY07 microbial inoculum has obviously promoted the growth of southerm yew seedling.Inoculation is handled height of seedling, the leading thread of southerm yew seedling and is compared the growth that all has in various degree with CK.Wherein, height of seedling and leading thread have increased by 24.15% and 17.09% respectively than CK.
Table 1CLY07 bacterial strain is to the influence of southerm yew seedling growth
Annotate: P＜0.05, same column different rows lowercase representative significant difference inequality.
Claims (2)
1. bacillus cereus CLY07, its called after bacillus cereus (Bacillus cereus) of classifying has been preserved in Chinese typical culture collection center C CTCC, deposit number: CCTCC NO:M 2010157, preservation date: on June 24th, 2010.
2. the application of the described bacillus cereus CLY07 of claim 1 in promoting the growth of southerm yew seedling.
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Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN1417325A (en) * | 2002-12-14 | 2003-05-14 | 山西省农业科学院植物保护研究所 | Waxy bacillus strain and microbial bactericide producing process with the strain |
| KR20030063956A (en) * | 2002-01-24 | 2003-07-31 | 주식회사 프로바이오닉 | Bacillus cereus Probio-32 strain for controlling parasitic nematode and method of treating nematode infections in plants |
| CN101591623A (en) * | 2009-04-15 | 2009-12-02 | 洪永聪 | A kind of bacillus cereus strain and with the degrading enzyme and preparation method thereof of the pyrethroid pesticide of its preparation |
| CN101629147A (en) * | 2009-06-30 | 2010-01-20 | 南京林业大学 | Bacillus cereus and application thereof in promoting growth of poplars |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR20030063956A (en) * | 2002-01-24 | 2003-07-31 | 주식회사 프로바이오닉 | Bacillus cereus Probio-32 strain for controlling parasitic nematode and method of treating nematode infections in plants |
| CN1417325A (en) * | 2002-12-14 | 2003-05-14 | 山西省农业科学院植物保护研究所 | Waxy bacillus strain and microbial bactericide producing process with the strain |
| CN101591623A (en) * | 2009-04-15 | 2009-12-02 | 洪永聪 | A kind of bacillus cereus strain and with the degrading enzyme and preparation method thereof of the pyrethroid pesticide of its preparation |
| CN101629147A (en) * | 2009-06-30 | 2010-01-20 | 南京林业大学 | Bacillus cereus and application thereof in promoting growth of poplars |
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