CN101709217A - Pseudomonas fluorescens CKD18 and application thereof - Google Patents
Pseudomonas fluorescens CKD18 and application thereof Download PDFInfo
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- CN101709217A CN101709217A CN200910241481A CN200910241481A CN101709217A CN 101709217 A CN101709217 A CN 101709217A CN 200910241481 A CN200910241481 A CN 200910241481A CN 200910241481 A CN200910241481 A CN 200910241481A CN 101709217 A CN101709217 A CN 101709217A
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Abstract
The invention provides Pseudomonas fluorescens CKD18, which has a strain preservation number of CGMCC No.3227. Experiments show that the strain has efficient stable phosphate-dissolving capability and can promote the P, N and K absorption of plants so as to promote plant growth. The strain can be made into various conditioners or bio-fertilizers and is highly worthy to be applied.
Description
Technical field
The present invention relates to new bacterial strain of a kind of microorganism and application thereof, be specifically related to a fluorescent pseudomonads and at activating soil indissoluble phosphorus with promote plant the application in aspect the absorption of P, N, K element and the growth thereof.
Background technology
Phosphorus is one of essential main nutrient elements of plant.Total phosphorus content in China's soil is considerable, but the phosphorus more than 95% exists with invalid forms such as stable aluminosilicate and phosphatic rock, plant is difficult to directly utilize, and can be on the low side by the available phosphorus content that plant absorbing is utilized, and therefore most farm crop and forest all will impose phosphate fertilizer.Yet the phosphate fertilizer this season utilization ratio that applies is only for 10%-25%, has at least the phosphorus of 70%-90% to enter soil and forms the solid form that is absorbed by crop for being difficult to.Simultaneously, long-term application phosphate fertilizer can reduce soil to the plain absorption of phosphorus, and the possibility of soil phosphorus validity and loss is increased; The excessive of phosphate fertilizer applies the decline that causes other constituent content of crop and biological effectiveness, thereby brings risk to HUMAN HEALTH.Therefore, improving the utilization ratio of phosphate fertilizer, improve the activation and the utilization of soil phosphorus, is the important focus of current research of agricultural science.
The circulation of nature phosphorus does not exchange with atmospheric sphere, and is therefore not only open but also have a deposition.Microorganism has middle cardiac status in natural phosphorus circulation, cyclic oxidation and Degradation by phosphorus compound realize this circulation.East wild arteries and veins emerging to studies confirm that of Dian Chi, Yunnan dissolving, conversion, migration, gathering and the sedimentary effect of phosphorus-solubilizing bacteria to phosphorus.From earlier 1900s, Stalstrom and Sackett find that some insoluble phosphoric acid salt of script and natural Rock Phosphate (72Min BPL) can have been carried out a large amount of researchs relevant for molten phosphorus microorganism by since the utilization that some bacteriums are dissolved.Gerretsen discovers that the plant dry weight that grows in the unsterilised soil increases 72-188% than the plant that grows in the sterile soil, and inhaling the phosphorus amount has increased 70-340%.
Microorganism with phosphorus decomposing ability comprises bacterium, fungi and actinomycetes, has a large amount of phosphorus bacteria fertilizers around soil and root system of plant, and wherein the quantity of phosphorus bacteria fertilizer is higher than the quantity of phosphorus bacteria fertilizer in the non-rhizosphere soil around the root system.Phosphorus bacteria fertilizer quantitatively wants many more than fungi, approximately is 2-150 times of fungi quantity.The bacterium of the molten phosphorus effect of tool of report mainly contains Bacillus (Bacillus), Rhodopseudomonas (Pseudomonas) at present, Escherichia (Escherichia), erwinia (Erwinia), Agrobacterium (Agrobacterium), root nodule bacterium (Bradyrhizobium), serratia (Serratia), Flavobacterium (Flavobacterium), enterobacteria belongs to (Enterbacte), micrococcus sp (Micrococcus), Azotobacter (Azotobacter), salmonella (Salmonella), chromobacterium (Chromobacterium), Alcaligenes (Alcaligenes), genus arthrobacter (Arihrobacter) and many Thiobacillus (Thiobacillus) etc.Can be divided into molten inorganic phosphorus bacterium and organophosphorus bacterium according to the kind of bacterolysis phosphorus, can decompose inorganic phosphorus (tricalcium phosphate, hydroxyapatite, Rock Phosphate (72Min BPL) etc.) Pseudomonas and mainly contain pseudomonas (Pseudomonas), Bacillus (Bacillus), rhizobium (Rhizobium), Achromobacterium (Achromobacter), Agrobacterium (Agrobacterium), micrococci (Microccocus), Aereobacter, Flavobacterium (Flavobacterium) and erwinia (Erwinia.) etc.The solubilized organophosphorus rhizobium (Rhizobium) arranged, enterobacteria (Enterobacter), serratia (Serratia), citric acid Pseudomonas (Citrobacter), proteus (Proteus), klebsiella (Klebsiella), false unit cell (Pseudomonas), bacillus (Bacillus).But simultaneously some bacterium can molten inorganic phosphorus again can dissolved organic phosphorus, as pseudomonas (Pseudomonas), bacillus (Bacillus), rhizobium (Rhizobium) etc.Molten phosphorus Mycophyta mainly contains Penicillium notatum (Penicillium), Aspergillus (Aspergillus), Rhizopus (Rhizopus), sickle-like bacteria (Fusarium), microsolerotium bacterium (Sclerotium) etc.Research at present focuses mostly in Penicillium (Penicillium), Aspergillus (Aspergillus) and Rhizopus (Rhizopus).Studies show that in numerous molten phosphorus microorganisms, the aspergillar activity is the highest.
There is number of mechanisms in the phosphorus decomposing of microorganism, hitherto reported organic acid generation, hydrogen proton, protein, sequestering action, oxygenizement etc. are arranged.Be commonly considered as owing to the inorganic or organic acid of microorganism secretion, and be accompanied by NH
4+The proton of assimilation discharges.When phosphorus bacteria fertilizer is inoculated in neutrality or the alkaline soil, the acid of generation has reduced the pH value of rhizosphere, has facilitated the dissolving of calcium phosphate and hydroxyapatite.Existingly studies show that different microorganism phosphorus decomposing mechanism there are differences.
Molten phosphorus microorganism is except having the effect of dissolving soil and external source indissoluble phosphorus, can improve the effect of crop yield in addition, the mechanism of its volume increase is many-sided: by nutritional status, secretion plant growth hormones, the secretion microbiotic that improves soil, the growth that improves mechanism such as the tolerance promotion plant of plant.
The comparison that the applied research of external phosphate solubilizing microorganism is carried out early, The former Russian scholar Meng Jina is separated to a kind of phosphorus decomposing bacillus megaterium in nineteen thirty-five from soil, and, it is reported P in the soil of inoculation back in nineteen forty-seven mass production and be widely used in USSR (Union of Soviet Socialist Republics) and East Bloc
2O
5Improve more than 15%.Gerretsen is seeded in phosphorus-solubilizing bacteria in the sand that a large amount of ground phosphate rock exist, plant-growth better.The research of the molten phosphorus microorganism of China starts from the fifties.Nineteen fifty-five, Chen Tingwei etc. isolate the acid no spore-bearing bacillus of a kind of product (Bact.sp) and have stronger dissolving phosphoric acid DFP ability from the wheat rhizosphere soil of Beijing.Corn and the millet sand culture experiment of inoculating this bacterium show: dry weight of maize has increased 32-45%, and the millet dry weight has increased by 51%.1988, the separation of Heilongjiang Province Institute of Micro-biology obtains phosphorus-solubilizing bacteria aspergillus niger (Aspergillus niger) AP-2, be made into phosphobacterin, under test conditions, increase soil and push up available phosphorus contents 141.9mg/L, and produce the cigarette district in the Heilongjiang Province and carried out field test, obtain positive effect, can save part phosphate fertilizer, produced tangible economic benefit.Release the series product of the various combination formation of multiple genus bacillus the eighties again, comprised bacillus cereus (Bacillus cereus), bacillus brevis (Bacillu.brevi), bacillus firmus (Bacillus.firmus).Ceng Guangqin etc. make microbial inoculum to the HM0332 of separation screening voluntarily and HM483 phosphate-solubilizing bacteria and are manured into soil, and reach 6.2-19.8% through the wheat increase yield rate of this bacterium inoculation, and the wheat quality of improvement is arranged, and improve soil quick-effective phosphor content, the effect of fertilizing soil.The use of molten phosphorus microorganism often is not single, often combines use between the multiple phosphorus-solubilizing bacteria or with many bacterial strains of root nodule bacterium, vinelandii, and a large amount of research has also been carried out in the interaction of molten phosphorus microorganism and other microorganisms.Wang Guanghua etc. to molten phosphorus microbial ecological effect study, measure with the DGGE fingerprint spectrum method of 16SrDNA and find, molten phosphorus fungi is seeded in the soil, improved the diversity of soybean and corn rhizospheric microorganism.Rudresh tests in greenhouse and big field, with phosphorus-solubilizing bacteria, root nodule bacterium and Trichoderma simultaneous inoculation in soil, discover that the three unites the rate of emergence that cultivation can promote garbanzo, promote absorption, the plant height of nutrition to increase, tiller, plant weight is improved, and the root nodule number increases, output is improved.And the effect that three's mixing applies is higher than any single applying and the barren effect.Singh etc. have reported the effect of inoculated with VA M and molten phosphorus microorganism on the mung bean, and VAM, the independent inoculation of phosphorus-solubilizing bacteria can both improve seed output, can obtain higher output yield when the two co-inoculation.As seen, the bacterium of efficient dissolving indissoluble phosphorus, for the exploitation of plant growth-promoting microorganism species and bio-feritlizer provides new microorganism resource, reduce the use of chemical fertilizer, improve the content of available phosphorus in the soil, improve edatope, reduce environmental pollution, in the hope of can be better being the agriculture production service, for the Sustainable development of agricultural contributes.Databases such as State Intellectual Property Office's patent retrieval database and United States Patent (USP), European patent do not find that as yet Pseudomonas fluorescens is with the application of efficient molten phosphorus microorganism on soybean and corn crop by retrieval.
Summary of the invention
The object of the present invention is to provide the new bacterial strain of a fluorescent pseudomonads and at activating soil indissoluble phosphorus with promote plant to the application in aspect the absorption of P, N, K element and the growth thereof.
Bacterial strain of the present invention is to separate the pseudomonas fluorescens CKD 18 that obtains from Huantai County, Shandong Province wheat experiment Tanaka.This bacterial strain on August 17th, 2009 at China Committee for Culture Collection of Microorganisms common micro-organisms center (address: No. 3, A, DaTun Road, Chaoyang District, BeiJing City, Institute of Microorganism, Academia Sinica, postcode 100101) preservation, classification called after Pseudomonas fluorescens (Pseudomonas fluorescens) CKD18, preserving number is CGMCCNo.3227.
CKD18 specifically separates by the following method and obtains: Tanaka gathers soil sample from the wheat experiment, the soil of getting 5g is dissolved in the triangular flask of 45ml stroke-physiological saline solution, static 1min behind 28 ℃, 120rpm vibration 15min, the supernatant liquor of getting 100 μ l joins in the Eppendorf pipe that 900 μ l physiological saline are housed, and carries out the continuous gradient dilution.Get above soil suspension and 10 respectively
-2, 10
-3, 10
-4Times soil suspension diluent 100 μ l are at NA substratum (extractum carnis 5g/L, peptone 10g/L, NaCl 5g/L, agar 15g/L) even coated plate on the flat board dries up in the Bechtop, each concentration repeats 3 times, cultivate 36h in 28 ℃ of incubators, behind the single bacterium colony line of aseptic toothpick picking bacterium purifying, be inoculated into Pikovsaia ' s Medium (PKO) substratum (glucose 10.0g, Yeast Extract 0.50g, Ca
3(PO
4) 25.0g, (NH
4)
2SO
40.5g, KCl 0.2g, MgCl
20.1g, MnSO
40.1mg, FeSO
40.1mg, agar 12-14.0g, H
2O 1000ml, PH 7.0-7.2,121 ℃ of sterilization 30min) in the flat board, cultivate 7d in 28 ℃ of constant temperature culture carton upside downs, observe molten phosphorus circle whether occurs, judge whether the molten phosphorus ability of tool, and measure the size of molten phosphorus circle, thereby phosphorus-solubilizing bacteria has been carried out preliminary screening; Detection to the tricalcium phosphate dissolving power is carried out in the phosphorus-solubilizing bacteria strain that obtains again, and CKD18 is 395.60mgL
-1, finishing screen is selected the good bacterial strain CKD18 of proterties (numbering CGMCC No.3227).
Morphological specificity, physio-biochemical characteristics, the interpretation of result of 16S rDNA sequencing of comprehensive Pseudomonas fluorescens are accredited as pseudomonas fluorescens CKD 18 with it.
Show that in the molten phosphorus effect detection test of the molten phosphorus effect of flat board and phosphoric acid three calcium solutions phosphorus-solubilizing bacteria strain CKD18 has stable, molten phosphorus ability efficiently; Absorb P, N, K element and promote in the plant-growth effect detection the molten phosphorus effect of soil and potted plant, CKD18 has improved the absorption to P, N, K element of soybean, corn, and has promoted the growth of plant.
Those skilled in the art are easy to expect bacterial strain fermentation liquor of the present invention or bacteria suspension are used for field crops as the effective constituent of bacteria agent.
The present invention also provides the CKD18 fermentation culture method, comprises steps such as actication of culture, seed liquor cultivation, ferment tank, specifically comprises the steps:
1, actication of culture
Use beef-protein medium, culture medium prescription is: extractum carnis 3.0g/l, peptone 10.0g/l, NaCl 5.0g/l, agar 12g/l.CKD18 is inoculated on the beef-protein medium inclined-plane, cultivates 48h for 28 ℃.
2, culture of seed liquid
Seed culture LB substratum, the prescription of LB substratum: peptone 10.0g/l, NaCl10.0g/l, Yeast Extract 5.0g/l, pH value are 7.0-7.2.The CKD18 that activation is good is mixed with 10 with physiological saline
8The bacteria suspension of cfu/ml, the inoculum size with 0.1% are inoculated in the LB liquid nutrient medium, and 28 ℃ of shaking table concussions are cultivated, and rotating speed is 180-200rpm, and incubation time is 40-48h.
3, ferment tank
Fermention medium consists of: glucose 10-12g/l, and corn steep liquor 3-5g/l, (NH4)
2SO
43-5g/l, KH
2PO
41-1.2g, MgSO
47H
2O 0.5-1.0g/l, CaCO
31-1.2g/l the pH value is 7.0-7.5.Per minute air flow volume), tank pressure 1.5-2.0F/cm2 cultured CKD18 seed liquor is inserted in the fermentor tank with 2% inoculum size, and 28 ℃, stirring velocity is 180rpm, and air flow is 1: 0.6-0.8 (fermentating liquid volume:.Fermentation 48h bacterium amount reaches 8-10 * 10
8Cfu/ml obtains the pseudomonas fluorescens CKD 18 fermented liquid.
Pseudomonas fluorescens CKD 18 of the present invention is at activating soil indissoluble phosphorus and promote plant to have very high research and using value in aspect the absorption of P, N, K element and the plant-growth thereof.
Description of drawings
Fig. 1 is the gel electrophoresis picture that the 16S rDNA of CKD18 carries out pcr amplification product, wherein, and 1, Marker; 2, blank; 3, the CKD18 genome is the 16SrDNA fragment of template amplification.
Fig. 2 is the phylogenetic tree according to the CKD18 of 16SrDNA sequence homology structure.
Fig. 3 is the dull and stereotyped molten phosphorus design sketch of the PKO of CKD18.
What Fig. 4 showed is the trend map that available phosphorus concentration changes with the CKD18 incubation time in the solution.
Embodiment
Following examples further specify content of the present invention, but should not be construed as limitation of the present invention.Without departing from the spirit and substance of the case in the present invention, modification or replacement to the inventive method, step or condition are done all belong to scope of the present invention.
If do not specialize the conventional means that used technique means is well known to those skilled in the art among the embodiment.
The evaluation of embodiment 1CKD18
Morphological specificity, physio-biochemical characteristics, the 16S rDNA sequencing result of comprehensive CKD18 bacterial strain are accredited as Pseudomonas fluorescens with it.Concrete qualification result is as follows:
1. physiological and biochemical property
The pseudomonas fluorescens CKD 18 physio-biochemical characteristics are as shown in table 1: Gram-negative bacteria, strict aerobic, gelatine liquefication, nitrate reduction, urase, Citrate trianion utilization, arginine dihydrolase, lecithinase, oxydase, H2S produce negative, and starch hydrolysis, denitrification, lipase, VP measure and all become positive; Can utilize fructose as carbon source, can not utilize sucrose, tartrate, glycine etc. as carbon source.
Table 1CKD18 physiological and biochemical property is measured project
+, positive reaction;-, negative reaction;
3,16S rDNA sequential analysis
Method is: the genome that extracts CKD18, with 16S amplification universal primer 8F:5`-CGGGATCCAGAGTTTGATCCTGGCTCAGAACGAACGCT-3` and 1506R:5`-CGGGATCCTACGGCTACCTTGTTACGACTTCACCCC-3`, in following PCR system, increase: 25 μ L amplification systems, each 0.25 μ L of 10mM primer, 10mM dNTP, 5U/ μ LTaqDNA polysaccharase (TaKaRa), 1 * PCR buffer (TaKaRa), 10ng/ μ L genomic dna is as template.The PCR reaction conditions is: 94 ℃ of pre-sex change of 5min, and 94 ℃ of 40S sex change, 56 ℃ of 40S annealing, 72 ℃ of 1min extend, and 30 circulations are fully extended 10min for back 72 ℃.Pcr amplification product is with 1% agarose gel electrophoresis analysis, reclaims the T carrier pMD18-T that the back connects TaKaRa company with the purification kit Gel Extraction Kit purifying of OMEGA company.Check order with ABI 3730 dna sequencing instrument in Invitrogen company, the result is shown in subordinate list.Utilize Blast software in GenBank, to carry out the homology comparison with other 16SrDNA sequence.The homology of the false unit cell of CKD18 and fluorescence is 99%.Simultaneously,, select close sequence, grow the tree (see figure 2) with the phyletic evolution of MEGA version 4 software building CKD18 according to above comparison result.
The phyletic evolution that comprehensive thalli morphology, physiological and biochemical property and 16Sr determined dna sequence and close sequence thereof are set up is grown tree, CKD18 is accredited as: Pseudomonas fluorescens (Pseudomonas fluorescens).
The dull and stereotyped molten phosphorus effect detection of embodiment 2CKD18 bacterial strain
Behind the single bacterium colony line of aseptic toothpick picking bacterium CKD18 purifying, be inoculated into Pikovsaia ' s Medium (PKO) solid medium (glucose 10.0g, yeast extract 0.50g, Ca
3(PO
4) 25.0g, (NH
4) 2SO
40.5g, KCl 0.2g, MgCl
20.1g, MnSO
40.1mg, FeSO
40.1mg, agar 12.0g, H
2O 1000ml, PH7.0-7.2,121 ℃ of sterilization 30min) in the flat board, cultivate 7d in 28 ℃ of constant temperature culture carton upside downs, measuring the molten phosphorus loop diameter of CKD18 is 16mm, it is stable that molten phosphorus ability keeps.(see figure 3)
Molten phosphorus effect detection in the embodiment 3CKD18 bacterial strain solution
With the CKD18 inoculation in the LB liquid medium, cultivate 48h in 170r/min, 28 ℃ of shaking tables, get bacterium liquid 50 μ l and be inoculated in the PKO liquid nutrient medium, in 28 ℃, 170r/min shaking table, shake training 10 days, with nutrient solution 12000r/min, 4 ℃ of centrifugal 5min get supernatant liquor 100 μ L (phosphorous 5-25 μ g), put into the 50ml volumetric flask, be diluted with water to about 30ml, add 2 of dinitrophenol indicator, drip 4mol/LNaOH and just transferred yellow to, add 2mol/L (1/2H again until solution
2SO
4) 1 droplet, the yellow of solution is just decorporated.Add the anti-developer of 5.00ml molybdenum antimony, the water constant volume fully shakes up.Room temperature be higher than 15 ℃ locate to place 30min after, survey at 882nm wavelength place with 1cm optical path cuvette and to read absorbancy, be reference liquid with the blank solution, regulate spectrophotometric zero point.
Accurately draw phosphorus standard operation solution (Cp=5mgL
-1) 0,0.50,1.00,2.00,4.00,6.00,8.00ml (in this standard serial solution the concentration of corresponding phosphorus be followed successively by 0,0.05,0.10,0.20,0.40,0.60,0.80mgL
-1P), put into the 50ml volumetric flask respectively, add water to about 30ml, add not inoculation culture liquid supernatant liquor 100 μ l more respectively, the pH of the same regulator solution and colour developing then, serial solution absorbency is read in survey, draws working curve, draws available phosphorus concentration value in the sample solution according to the typical curve of drawing.
Experiment shows that CKD18 is in the process of cultivating, and available phosphorus concentration is slow straight line rising type in the solution.(see figure 4)
The molten phosphorus ability of embodiment 4CKD18 bacterial strain soil detects
Test used soil from Beijing China Agricultural University test base, content of tatal phosphorus 0.019%, total nitrogen content 0.30%, full potassium content 3.10%, available phosphorus content 5.03mg/kg, organic carbon content 22.74g/kg, organic content 3.92mg/kg.Test is with native: soil: the peat composed of rotten mosses: vermiculite (V: V: V)=1: 1: 1; With the ground phosphate rock crushing screening (＜0.018mm), press 5.0g: 1000g (ground phosphate rock: soil) amount with native mixing.Soil disinfection adopts moist heat sterilization, 121 ℃ of sterilization 30min.Test design such as table 2.4 repetitions are established in each processing, every basin 250g soil, and inoculation bacterium amount is 50ml bacterium liquid/Kg soil, blank then applies sterilization LB nutrient solution 50ml/Kg soil, totally 40 basins, sampling and measuring soil available phosphorus behind 42d.
The molten phosphorus ability test design of table 2CKD18 soil
*: standard deviation;
#Variance analysis, p=0.05
Test-results is as shown in table 3, and bacterial classification inoculation not adding outside phosphorus source and adding in the soil in outside phosphorus source, is compared with not inoculating blank, and available phosphorus concentration all increases in the soil.
In sterile soil, do not add outside phosphorus source and handle down, inoculation CKD18 available phosphorus increases by 4.5%, does not reach conspicuous level.Under the processing of adding outside phosphorus source, the soil available phosphorus increase of inoculation phosphorus bacteria fertilizer reaches utmost point conspicuous level, and available phosphorus increases by 27.5%, illustrates that this bacterial strain has good molten phosphorus effect in sterile soil.
In non-sterile soil, do not apply outside phosphorus source and handle down, the processing available phosphorus of inoculation bacterium CKD18 is than blank height, and available phosphorus has improved 5.0%, illustrate that the CKD18 bacterial strain is good with other microorganisms symbiosiss in soil, and this symbiosis does not influence the performance of its molten phosphorus ability.Add in the non-sterile soil under the situation in outside phosphorus source, available phosphorus concentration and blank difference were extremely remarkable under inoculation CKD18 handled, and the processing available phosphorus of inoculation CKD18 has increased by 37.2%.
Embodiment 5 short giving birth to and molten phosphorus effect measuring
Soil adopts sterilization soil, seed 30%H
2O
2Surface sterilization, the back places 28 ℃ of incubators to carry out vernalization on moistening gauze, and corn 48h prior to seeding carries out vernalization, and soybean 24h vernalization prior to seeding gets final product.Greenhouse pot culture is handled and is selected for use corn (Huan rich 16) and soybean (Xindou No.1) for the test seed, and the processing that is added with CKD18 bacterium liquid and ground phosphate rock in space management and the soil is set.Each handles 4 repetitions, every basin 1000g soil, and inoculation bacterium amount is 50ml bacterium liquid/Kg soil, blank then sterilize LB 50ml/Kg soil, totally 24 basins.Behind 42d, receive the seedling test sample.Measure the plant dry weight and fresh weight of plant seedlings, oven dry disappears and boils the full nitrogen of back measuring plants, full phosphorus, full potassium content.
The potted plant soil available phosphorus of table 4 changes (available phosphorus mg/kg)
*: standard deviation;
#Variance analysis, p=0.05
As shown in table 4 behind the plantation plant to the solubility test result of soil available phosphorus, compare with blank, available phosphorus content increases to some extent in the potted plant soil in inoculation bacterium and the outside phosphorus of interpolation source, and the potted plant available phosphorus of corn of inoculation CKD18 is compared with blank and improved 23%, and soybean has improved 18%.
Efficient phosphorus-solubilizing bacteria CKD18 is to the dry weight of corn and the influence of fresh weight (g/ basin) in table 5 inoculation
*: standard deviation;
#Variance analysis, p=0.05
Efficient phosphorus-solubilizing bacteria CKD18 is to dry weight of soybean and the influence of fresh weight (g/ basin) in table 6 inoculation
*: standard deviation;
#Variance analysis, p=0.05
After inoculating efficient phosphorus-solubilizing bacteria CKD18, can effectively increase the ground and the subterranean dry weight and fresh weight of plant seedlings of corn.The result is as shown in table 5: the inoculation of phosphorus-solubilizing bacteria CKD18 has increased corn field top fresh weight and dry weight, CKD18 makes corn field top fresh weight increase by 23.9%, underground fresh weight has increased by 47.0%, and the over-ground part dry weight has increased by 17.1%, and the underground part dry weight has increased by 28%.The inoculation of CKD18 bacterial strain to increase on the soybeans, the underground part dry weight and fresh weight of plant seedlings has the trend of increase.
The efficient phosphorus-solubilizing bacteria CKD18 of table 7 inoculation is to the influence (mg/ basin) of corn NPK
The efficient phosphorus-solubilizing bacteria CKD18 of table 8 inoculation is to the influence (mg/ basin) of soybean NPK
After inoculating efficient phosphorus-solubilizing bacteria CKD18, can significantly increase the absorption of corn, also increase the absorption of corn simultaneously nitrogen and potassium to phosphorus; Can effectively increase the absorption of soybean, but increase not obvious the absorption of soybean nitrogen and potassium to phosphorus.The result is shown in table 7 and table 8: no matter be corn or soybean, the full phosphorus of plant all increases, and wherein corn phosphorus is absorbed promoter action tool utmost point significance, and the corn content of tatal phosphorus has increased by 43.8%, and the soybean content of tatal phosphorus has increased by 7.6%.In corn test, CKD18 has promoted the absorption of corn to the N element, reaches utmost point conspicuous level with blank difference, has improved 27.6% with not inoculating to compare; To the absorption of K element, the processing of inoculation CKD18 has increased by 35.6% than blank.In soy bean test, CKD18 has promoted the absorption of soybean to the K element, has increased by 5.2% compared with the control.
Sequence table
＜110〉China Agricultural University
＜120〉Pseudomonas fluorescens CKD 18 and application thereof
<130>KHP09113811.4
<160>3
<170>PatentIn?version?3.5
<210>1
<211>38
<212>DNA
＜213〉artificial sequence
<400>1
cgggatccag?agtttgatcc?tggctcagaa?cgaacgct 38
<210>2
<211>36
<212>DNA
＜213〉artificial sequence
<400>2
cgggatccta?cggctacctt?gttacgactt?cacccc 36
<210>3
<211>1451
<212>DNA
<213>Pseudomonas?fluorescens
<400>3
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ttcagcggcg?gacgggtgag?taatacctag?gaatctgcct?ggtagtgggg?gacaacgttt 120
cgaaaggaac?gctaataccg?catacgccct?acgggggaaa?gcaggggacc?ttcgggcctt 180
gcgctatcag?atgagcctag?gtcggattag?ctagttggtg?aggtaatggc?tcaccaaggc 240
gacgatccgt?aactggtctg?agaggatgat?cagtcacact?ggaactgaga?cacggtccag 300
actcctacgg?gaggcagcag?tggggaatat?tggacaatgg?gcgaaagcct?gatccagcca 360
tgccgcgtgt?gtgaagaagg?tcttcggatt?gtaaagcact?ttaagttggg?aggaagggta 420
cttacctaat?acgtgagtat?tttgacgtta?ccgacagaat?aagcaccggc?taactctgtg 480
ccagcagccg?cggtaataca?gagggtgcaa?gcgttaatcg?gaattactgg?gcgtaaagcg 540
cgcgtaggtg?gttcgttaag?ttggatgtga?aatccccggg?ctcaacctgg?gaactgcatc 600
caaaactggc?gagctagagt?agggcagagg?gtggtggaat?ttcctgtgta?gcggtgaaat 660
gcgtagatat?aggaaggaac?accagtggcg?aaggcgacca?cctgggctca?tactgacact 720
gaggtgcgaa?agcgtgggga?gcaaacagga?ttagataccc?tggtagtcca?cgccgtaaac 780
gatgtcaact?agccgttgga?atccttgaga?ttttagtggc?gcagctaacg?cattaagttg 840
accgcctggg?gagtacggcc?gcaaggttaa?aactcaaatg?aattgacggg?ggcccgcaca 900
agcggtggag?catgtggttt?aattcgaagc?aacgcgaaga?accttaccag?gccttgacat 960
gcagagaact?ttccagagat?ggattggtgc?cttcgggaac?tctgacacag?gtgctgcatg 1020
gctgtcgtca?gctcgtgtcg?tgagatgttg?ggttaagtcc?cgtaacgagc?gcaacccttg 1080
tccttagtta?ccagcacgtt?atggtgggca?ctctaaggag?actgccggtg?acaaaccgga 1140
ggaaggtggg?gatgacgtca?agtcatcatg?gcccttacgg?cctgggctac?acacgtgcta 1200
caatggtcgg?tacagagggt?tgccaagccg?cgaggtggag?ctaatctcac?aaaaccgatc 1260
gtagtccgga?tcgcagtctg?caactcgact?gcgtgaagtc?ggaatcgcta?gtaatcgcga 1320
atcagaatgt?cgcggtgaat?acgttcccgg?gccttgtaca?caccgcccgt?cacaccatgg 1380
gagtgggttg?caccagaagt?agctagtcta?accttcggga?ggacggttac?cacggtgtga 1440
tcatgaccgt?t 1451
Claims (8)
- Pseudomonas fluorescens at activating soil indissoluble phosphorus, promote plant that P, N, K are absorbed or promote application in the plant-growth.
- 2. application as claimed in claim 1 is characterized in that, described Pseudomonas fluorescens is Pseudomonas fluorescens CKD 18.
- 3. the microbial inoculum of an activating soil indissoluble phosphorus, it contains Pseudomonas fluorescens.
- 4. require 3 described microbial inoculums as claim, it is characterized in that, described Pseudomonas fluorescens is Pseudomonas fluorescens CKD 18.
- 5. the soil conditioner that contains claim 3 or 4 described microbial inoculums.
- 6. the fertilizer that contains claim 3 or 4 described microbial inoculums.
- 7. fertilizer as claimed in claim 6, it is a bio-fertilizer.
- 8. Pseudomonas fluorescens (Pseudomonas fluorescens) CKD18, deposit number is CGMCC No.3227.
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| CN2009102414814A CN101709217B (en) | 2009-12-03 | 2009-12-03 | Pseudomonas fluorescens CKD18 and application thereof |
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