CN114806924B - Achromobacter denitrificans and application thereof - Google Patents
Achromobacter denitrificans and application thereof Download PDFInfo
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- CN114806924B CN114806924B CN202210249420.8A CN202210249420A CN114806924B CN 114806924 B CN114806924 B CN 114806924B CN 202210249420 A CN202210249420 A CN 202210249420A CN 114806924 B CN114806924 B CN 114806924B
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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
- C05G3/90—Mixtures of one or more fertilisers with additives not having a specially fertilising activity for affecting the nitrification of ammonium compounds or urea in the soil
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- 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
- C12N1/20—Bacteria; Culture media therefor
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02W—CLIMATE CHANGE MITIGATION TECHNOLOGIES RELATED TO WASTEWATER TREATMENT OR WASTE MANAGEMENT
- Y02W30/00—Technologies for solid waste management
- Y02W30/40—Bio-organic fraction processing; Production of fertilisers from the organic fraction of waste or refuse
Abstract
The invention discloses a denitrifying achromobacter and application thereof. The preservation number of the denitrifying achromobacter (Achromobacter denitrificans) YSQ030 is CGMCC NO.22850. The denitrifying achromobacter nitrous oxide obtained by separation has strong emission reduction effect, has stronger adaptability in stress environment, can effectively promote the growth of crops, and can be widely applied to farmland soil such as rice, corn and the like.
Description
Technical Field
The invention belongs to the field of agricultural microbiological technology and environmental protection, and relates to a denitrifying achromobacter and application thereof.
Background
Nitrous oxide (N) 2 O), a greenhouse gas, can be transported to the stratosphere, resulting in ozone depletion and ozone depletion. Nitrous oxide is very low in atmospheric content, belongs to trace gases, but has long residence time in the atmosphere, and the heating potential (GWP) of single molecules on the scale of hundred years of nitrous oxide is CO 2 The warming effect of increasing nitrous oxide concentration on global climate has led to a tremendous increase in scientists by 265 times (IPCC, 2014)Attention is paid.
Agricultural sources are important sources of nitrous oxide, which are about 58% of the total national nitrous oxide emissions and grow year by year. The large investment of nitrogen fertilizer in agricultural production is one of the important reasons for the increase of the concentration of nitrous oxide in the atmosphere. The method for reducing the nitrous oxide in farmland soil mainly comprises fertilization management (including fertilization time and mode, fertilization amount, fertilizer type and the like), nitrification inhibitor use, scientific cultivation, biomass charcoal addition, inoculation of microorganisms with nitrous oxide emission reduction effect and the like. Compared with other methods, the nitrous oxide emission reduction is carried out by adding the microbial agent, so that the method is more economical and easy to operate.
At present, few microbial strains capable of reducing the nitrous oxide emission of soil are adopted, most of the microbial strains adopt nitrous oxide reducing bacteria, the effect is single, and only the effect of reducing the nitrous oxide emission can be realized.
Disclosure of Invention
In order to solve the defects that the microorganism strain with the effect of reducing the nitrous oxide emission of the soil is single in variety and single in action on farmland soil in the prior art, the invention provides the denitrifying achromobacter which can effectively realize the nitrous oxide emission reduction and improve the crop yield.
The invention relates to a denitrifying achromobacter, which is classified and named as denitrifying achromobacter @Achromobacter denitrificans) The strain is named YSQ030, and the preservation number is CGMCC NO.22850. The biochemical identification results are shown in Table 1.
The invention provides application of the denitrifying achromobacter in preparation of microbial agents.
Specifically, the microbial agent is prepared by the following method: and (3) inoculating the denitrifying achromobacter into a fermentation culture medium with the pH of 7.2-7.5, and culturing at 26+/-2 ℃ until the viable count reaches 2 hundred million/ml or more than 2 hundred million/g, thereby obtaining the microbial agent.
Wherein the fermentation medium is peptone 5.0 g/L, beef extract powder 3.0 g/L, sodium nitrate 0.3 mM and sodium succinate 4.4 mM, and the solvent is water; the fermentation medium can also be 10.0 g/L tryptone, 5.0 g/L yeast extract powder, 10.0 g/L sodium chloride, 0.3 mM sodium nitrate and 4.4 mM sodium succinate, and the solvent is water; the fermentation medium can also be tryptone 10.0 g/L, maltose 5.0 g/L, sodium chloride 10.0 g/L, sodium nitrate 0.3 mM and sodium succinate 4.4 mM, and the solvent is water; the fermentation medium can also be 10.0 g/L soybean peptone, 10.0 g/L sucrose, 5.0 g/L sodium chloride, 0.3. 0.3 mM sodium nitrate and 4.4. 4.4 mM sodium succinate, and the solvent is water.
The invention also provides application of the denitrifying achromobacter in preparation of a biological organic fertilizer.
Specifically, the biological organic fertilizer is prepared by inoculating the microbial agent to the organic fertilizer; wherein the inoculation proportion of the microbial agent is 1 milliliter of microbial agent inoculated per gram of organic fertilizer.
The invention also provides application of the denitrifying achromobacter in reducing farmland greenhouse gas nitrous oxide emission and application of the denitrifying achromobacter in improving soil microbial community structures.
The farmland or soil is farmland of rice or corn.
The beneficial effects of the invention are as follows: the microbial strain is obtained by separating from the rhizosphere soil of the pond reed, is biochemically identified to be the denitrifying achromobacter, has strong nitrogen oxide emission reduction effect, has strong adaptability in a high-salt and alkaline environment, can effectively promote the growth of crops, and can be widely applied to farmland soil such as rice, corn and the like.
The strain of the invention has a preservation date of 2021, 7 and 8 days and a preservation number of CGMCC NO.22850. The classification is named: achromobacter denitrificans @Achromobacter denitrificans) YSQ030. The preservation unit is China general microbiological culture Collection center, and the address is the post code 100101 of the microbiological institute of China academy of sciences.
Drawings
FIG. 1 is a phenotypic characteristic of Achromobacter denitrificans YSQ 030;
FIG. 2 is a physiological and biochemical characterization of Achromobacter denitrificans YSQ 030;
FIG. 3 is a representation of the growth of Achromobacter denitrificans YSQ030 at different pH values;
FIG. 4 is a representation of the growth of Achromobacter denitrificans YSQ030 at different salinity;
FIG. 5 is a graph showing the effect of Achromobacter denitrificans YSQ030 on nitrous oxide emission flux when mixed with organic fertilizer (without soil addition);
FIG. 6 shows the effect of Achromobacter denitrificans YSQ030 on cumulative nitrous oxide emissions from rice soil under microcosmic conditions;
FIG. 7 shows the effect of Achromobacter denitrificans YSQ030 on cumulative nitrous oxide emissions from corn soil under microcosmic conditions.
Detailed Description
The invention will be better understood from the following examples. However, it will be readily appreciated by those skilled in the art that the description of the embodiments is provided for illustration only and should not limit the invention as described in detail in the claims.
EXAMPLE 1 isolation and screening of Achromobacter denitrificans YSQ030
Collecting the rhizosphere soil of reed in a certain pond in new areas of Nanjing, jiangbei province, weighing 10 parts g, and transferring to an ultra-clean workbench for operation. Placing the soil sample into a conical flask containing 90 mL of 0.87% NaCl solution, and shaking at 28deg.C and 200 r/min for 1 h to obtain a dilution of 10 -1 Is a soil suspension of (a); take 10 -1 Is put into a conical flask containing 90 mL of 0.87% NaCl solution, and is evenly mixed to prepare the soil suspension 10 with the dilution degree of 10 -2 Is a soil suspension of (a); and so on. Respectively preparing the components with dilution degree of 10 -3 、10 -4 、10 -5 Is a soil suspension of (a). Then respectively taking out 10 by using a pipettor -3 、10 -4 、10 -5 100. Mu.L of the dilution of (A) was applied to a nitrogen-fixing solid medium (KH-containing per liter) 2 PO 4 ，0.2 g；K 2 HPO 4 ，0.8 g；MgSO 4 ·7H 2 O，0.2 g；CaSO 4 ·2H 2 O，0.1 g；FeCl 3 Trace amount; na (Na) 2 Mo 4 ·2H 2 O, trace; yeast extract, 0.5 g; mannitol, 20g, agar, 15 g; pH7.2, water as solvent), each gradientThree replicates. And (3) culturing in an inverted mode in a biochemical incubator at the constant temperature of 28 ℃ for 2-4 d, and observing the growth condition of the strain every day. After the single colony appeared, a relatively large colony was picked up with an inoculating loop in an ultra clean bench and inoculated into 1/100 NBNS (peptone 5 g.L) -1 Beef soaked powder 3 g.L -1 ，0.3 mM NaNO 3 And 4 mM sodium succinate, pH 7.2-7.5, water as solvent) for 5-6 generations, and 30% glycerol is used for preserving at-80 ℃ for later use.
EXAMPLE 2 identification of Achromobacter denitrificans YSQ030
Culturing the colony on an NBNS agar culture medium, wherein the colony is dry and round, slightly raised, light yellow, moist, semitransparent, and neat and smooth in edge; bacterial film formation occurs when the liquid culture is stationary, and is shown in fig. 1, wherein a is morphological characteristics on a plate, and b is fermentation broth state of the liquid culture. Gram staining was negative, rod-like, spore-free, liquefied gelatin. By 16S rDNA sequence analysis (see nucleotide sequence SEQ ID NO: 1), homology to Achromobacter denitrificans after BLAST alignment in NCBI; the physiological and biochemical characteristics are shown in table 1 and fig. 2.
TABLE 1 physiological and biochemical Properties of Achromobacter denitrificans YSQ030
Example 3 analysis of the growth Capacity of Achromobacter denitrificans YSQ030 at different pH values
Achromobacter denitrificans YSQ030 strain solution with an OD value of 100 microliters was inoculated into 10 mL NBNS liquid medium with a pH of 4, 6, 8 and 10, respectively (volume ratio of about 1:100), shake-cultured at 26℃and 160 r/m for 6, 12 and 24 hours, and strain growth curves were drawn by measuring OD600 values with the NBNS liquid medium without adding strain as a control. Three replicates were set for each set of experiments and the results are shown in figure 3. As a result, it was found that Achromobacter denitrificans YSQ030 is most suitable for survival in a neutral environment, but has a good viability in an alkaline environment and a good tolerance in an alkaline environment.
Example 4 analysis of the growth Capacity of Achromobacter denitrificans YSQ030 under different salinity
Achromobacter denitrificans YSQ030 strain solutions with an OD of 100. Mu.l were inoculated into 10 mL NBNS liquid medium with a salinity of 0%,4%,8% and 10%, respectively (volume ratio: about 1:100), shake-cultured at 26℃and 160 r/m for 6, 12 and 24 hours, and strain growth curves were drawn by measuring OD600 values with respect to the NBNS liquid medium without adding strain as a control. Three replicates were set for each set of experiments and the results are shown in figure 4. The test result shows that the denitrifying achromobacter YSQ030 is more suitable for survival under a high-salt environment: achromobacter denitrificans YSQ030 has a growth ability far superior to other strains at salinity of 8% and 10%.
EXAMPLE 5 analysis of Nitrogen oxide emission reduction Effect of Achromobacter denitrificans YSQ030 when mixed with organic Fertilizer (without soil addition)
Preparing a bacterial suspension: single colony of denitrifying achromobacter is picked up and inoculated to NBNS (peptone 5 g.L) -1 Beef extract 3 g L -1 ，0.3 mM NaNO 3 And 4 mM sodium succinate, pH 7.2-7.5, water as solvent), and culturing at 28deg.C in a shaker at 160 rpm for more than 24 hr. The OD600 = 1.0 was adjusted with NBNS broth. 20g of an organic fertilizer with a nitrogen content of 7% (in this example, the organic fertilizer is selected from commercial organic fertilizers of the "Di Yi" brand produced by Nanjing Ming's fertilizer Co., ltd.) and 20mLOD of Achromobacter denitrificans YSQ030 strain liquid with a value of 1 are placed in a culture flask. Keeping the organic fertilizer in a flooded state. The organic fertilizer-bacterial liquid mixture was placed in a glass bottle (500 ml) covered with a plastic cap having a number of small holes and then incubated at 26 ℃. To measure nitrous oxide flux, 1 hour after closing the cap by a plastic cap of a silicon septum, 1.0ml headspace gas was sampled from the bottle using a glass syringe, and then analyzed by a gas chromatography electron capture detector (ECD-GC). Control comparisons were made with sterile NBNS broth without inoculum (CK group). The results are shown in FIG. 5. The denitrifying achromobacter YSQ030 has an emission reduction effect on nitrous oxide when mixed with an organic fertilizer (without adding soil). As can be seen from FIG. 5, inoculation with Achromobacter denitrificans YSQ030 (YSQ 030 group) significantly reduced organic fertilizer oxidationCumulative emissions of nitrous oxides; the same species of Achromobacter denitrificans (LS 20 and LS 23) respectively reduced the cumulative emission of nitrous oxide in the organic fertilizer by 34% and 22% as compared with the control, while Achromobacter denitrificans YSQ030 reduced the cumulative emission of nitrous oxide in the organic fertilizer by more than 37% as compared with the control.
Example 6 analysis of reduction of nitrous oxide emissions from soil by Achromobacter denitrificans YSQ030 under microcosmic conditions
Test soil: collecting soil on the surface layer of Jiangsu Yixing vegetable, naturally airing, and sieving with a 2 mm sieve for later use. The soil properties of the surface layer of Jiangsu Yixing vegetable land are as follows: pH:7.69; conductivity: 140.5 μs/cm; TC:37.98mg/g (per gram of air-dried soil); TN:0.87mg/g (per gram of air-dried soil); C/N:46.72. preparing a bacterial suspension: single colonies of Achromobacter denitrificans were picked up and inoculated into NBNS (peptone 5 gL) -1 Beef extract 3 gL -1 ，0.3 mM NaNO 3 And 4 mM sodium succinate, pH 7.2-7.5, water as solvent), and culturing at 28deg.C in a shaker at 160 rpm for more than 24 hr. The OD600 = 1.0 was adjusted with NBNS broth. Microcosm inoculation test: weighing 100 g air-dried soil into a 500mL culture bottle, adding a mixture of 5 mL bacterial liquid and 5g of organic fertilizer, and fully and uniformly mixing (YSQ 030 groups); control 2 was added with an equal amount of a mixture of NBNS liquid medium and 5g of organic fertilizer (NBNS group); proper amount of sterile water is added to ensure that the soil moisture reaches 80% of the maximum field water holding capacity; finally, the culture is carried out at 26 ℃ in dark. Nitrous oxide collection: and (3) adopting a sealed culture gas sampling method, sealing for 1 hour, collecting a gas sample at the top of a culture bottle, analyzing the concentration of nitrous oxide by utilizing gas chromatography containing an ECD detector, and calculating the nitrous oxide emission flux and the accumulated emission quantity. The results are shown in FIG. 6. Is the influence of the denitrifying achromobacter YSQ030 on the nitrous oxide emission flux of paddy soil under the microcosmic condition. As can be seen from fig. 6, inoculation of achromobacter denitrificans YSQ030 (YSQ 030 group) significantly reduced cumulative discharge amount of nitrous oxide in paddy soil; the accumulated discharge of nitrous oxide in dry soil was reduced by approximately 30% and 15% respectively for the same species of Achromobacter denitrificans (LS 20 and LS 23) as compared to the control, while the nitrous oxide in dry soil was reduced by Achromobacter denitrificans YSQ030 as compared to the control (NBNS group)The cumulative emissions are approximately 50%.
Example 7 analysis of reduction of nitrous oxide emissions from corn soil by Achromobacter denitrificans YSQ030 under microcosmic conditions
Test soil: collecting soil on the surface of North China plain in Shandong, naturally air-drying, and sieving with a 2 mm sieve for later use. The corn soil properties in Shandong Texas are as follows: pH:7.33; conductivity: 163.35 μs/cm; TC:0.026g (per gram of air-dried soil); TN:0.00087g (per gram of air-dried soil); C/N:30.54. bacterial liquid preparation, microcosmic inoculation test and nitrous oxide collection were the same as in example 6. As shown in fig. 7, the results demonstrate that inoculation of achromobacter denitrificans YSQ030 (YSQ 030 group) significantly reduces cumulative discharge of nitrous oxide from corn soil; achromobacter denitrificans (LS 20 and LS 23) of the same species reduced the cumulative emissions of nitrous oxide from corn soil by 10% and 12%, respectively, as compared to the control, while Achromobacter denitrificans YSQ030 reduced the cumulative emissions of nitrous oxide from corn soil by 25.6% as compared to the control (NBNS group).
Example 8 analysis of Achromobacter denitrificans YSQ030 in Water culture experiments to promote Rice growth
Pre-breeding of rice seedlings: and (3) placing the preserved south japonica 46 rice seeds into a 50mL centrifuge tube, adding 2.5% NaClO solution, and shaking up and down for 10min to sterilize the surfaces of the rice seeds. Next, the seeds were rinsed 10 times with clean tap water to remove residual NaClO on the seed surface until no obvious sterilizing water (chlorine) smell was observed. 2 basins of clean seedling raising trays are selected, gauze with proper size is trimmed and placed on the upper layer of the seedling raising trays, sterilized rice seeds are uniformly sown on the gauze, and clean tap water is added until water is discharged by lightly pressing the gauze. When the rice seeds on the gauze begin to emit white buds, the rice seeds are continuously cultured by using 1/5 Hoagland nutrient solution. After accelerating germination for 10 days, selecting seedlings with uniform size and consistent growth vigor, wrapping the stem base parts of the seedlings with sponge, transplanting the seedlings into culture pots respectively, and culturing the seedlings for 3 days by using 1/10 Hoagland nutrient solution. Bacterial liquid treatment: pre-cultured rice seedlings were divided into 2 groups of 10 pots each, 5 seedlings per pot. The first group is used as control, NBNS culture solution with the same amount as bacterial solution is added, and the second group is inoculated with 10 5 CFU ml -1 Is a bacterial liquid of (a) a strain. The preparation method of the bacterial liquid is referred to in example 5. The culture medium is replaced every 3-4 days. Measurement of rice growth: harvesting at 14 days after treatment, measuring the plant height of the rice by using a ruler, measuring the fresh weights of the overground parts and the overground parts respectively, and calculating the root cap ratio. The specific results are shown in Table 2. As can be seen from table 2, compared with the control, the achromobacter denitrificans YSQ030 significantly improved the root fresh weight, the overground part fresh weight and the total fresh weight of the rice seedlings, and had no significant effect on the plant height and root cap ratio of the rice seedlings.
TABLE 2 Effect of Achromobacter denitrificans YSQ030 on rice growth
| Group of | Repeating | Plant height/cm | Fresh root weight/mg | Fresh weight of overground part/mg | Total fresh weight/mg | Root-to-crown ratio |
| Control | 1 | 18.8 | 53 | 53 | 106 | 1.00 |
| 2 | 13.4 | 24 | 28 | 52 | 0.86 | |
| 3 | 16.0 | 28 | 36 | 64 | 0.78 | |
| 4 | 18.7 | 72 | 59 | 131 | 1.22 | |
| 5 | 17.2 | 39 | 47 | 86 | 0.83 | |
| 6 | 16.1 | 49 | 47 | 96 | 1.04 | |
| 7 | 18.6 | 69 | 64 | 133 | 1.08 | |
| 8 | 17.3 | 55 | 43 | 98 | 1.28 | |
| 9 | 19.1 | 70 | 57 | 127 | 1.23 | |
| 10 | 19.2 | 97 | 53 | 150 | 1.83 | |
| Average value of | 17.4a | 56b | 49b | 104b | 1.14a | |
| YSQ030 | 1 | 18.9 | 83 | 76 | 159 | 1.09 |
| 2 | 17.9 | 84 | 78 | 162 | 1.08 | |
| 3 | 20.0 | 105 | 97 | 202 | 1.08 | |
| 4 | 18.6 | 71 | 73 | 144 | 0.97 | |
| 5 | 16.2 | 62 | 70 | 132 | 0.89 | |
| 6 | 15.4 | 49 | 52 | 101 | 0.94 | |
| 7 | 19.7 | 80 | 82 | 162 | 0.98 | |
| 8 | 17.4 | 78 | 69 | 147 | 1.13 | |
| 9 | 16.1 | 92 | 76 | 168 | 1.21 | |
| 10 | 14.3 | 61 | 49 | 110 | 1.24 | |
| Average value of | 17.5a | 77a | 72a | 149a | 1.06a | |
| Increase rate% | 0.1 | 37.6 | 48.3 | 42.6 | -7.2 |
Example 9 analysis of Achromobacter denitrificans YSQ030 to reduce discharge of nitrous oxide from soil of rice and promote growth of rice in field test
Field test site: nanjing Liuhe. The basic physicochemical properties of the six-in paddy soil are as follows: soil pH:5.9; 25.4. 25.4 g/kg of organic matters; total nitrogen 1.68 g/kg; available phosphorus 32 mg/kg; quick-acting potassium 108/mg/kg. The test began at 7 months 2020. Each cell size was experimentally 4 m ×5 m, stem width 0.6 m, guard line width 2.5 m, channel width 1 m. Base fertilizer: applying bio-organic fertilizer 2 kg of Achromobacter denitrificans YSQ030 (YSQ 030 group, mixed bacterial liquid, bacterial liquid and organic fertilizer same as example 6) and urea 0.35 kg (control group); tillering fertilizer: urea 0.33 kg was applied; jointing booting fertilizer: urea 0.11, kg was applied. When harvesting rice in season, the cell yield of the control group is reduced by 8059.5 kg ha -1 Cell yield reduced 9026.4 kg. Ha with the addition of Achromobacter denitrificans (group YSQ 030) -1 12.0% higher than the control group. The accumulated discharge amount of nitrous oxide in the whole growth period of rice is reduced by 20 percent.
The sequence table of the denitrifying achromobacter of the invention is as follows:
<110> university of Nanjing information engineering
<120> Achromobacter denitrificans and application thereof
<160> 1
<170> SIPOSequenceListing 1.0
<210> 1
<211> 1407
<212> DNA
<213> Achromobacter denitrificans (Achromobacter denitrificans)
<400> 1
CATGCAAGTC GAACGGCAGC ACGGACTTCG GTCTGGTGGC GAGTGGCGAA CGGGTGAGTA 60
ATGTATCGGA ACGTGCCTAG TAGCGGGGGA TAACTACGCG AAAGCGTAGC TAATACCGCA 120
TACGCCCTAC GGGGGAAAGC AGGGGATCGC AAGACCTTGC ACTATTAGAG CGGCCGATAT 180
CGGATTAGCT AGTTGGTGGG GTAACGGCTC ACCAAGGCGA CGATCCGTAG CTGGTTTGAG 240
AGGACGACCA GCCACACTGG GACTGAGACA CGGCCCAGAC TCCTACGGGA GGCAGCAGTG 300
GGGAATTTTG GACAATGGGG GAAACCCTGA TCCAGCCATC CCGCGTGTGC GATGAAGGCC 360
TTCGGGTTGT AAAGCACTTT TGGCAGGAAA GAAACGTTCC GGGGTTAATA CCCCGGGAAA 420
CTGACGGTAC CTGCAGAATA AGCACCGGGC TAACTACGTG CCAGCAGCCG CGGTAATACG 480
TAGGGTGCAA GCGTTAATCG GAATTACTGG GCGTAAAGCG TGCGCAGGCG GTTCGGAAAG 540
AAAGATGTGA AATCCCAGAG CTTAACTTTG GAACTGCATT TTTAACTACC GAGCTAGAGT 600
GTGTCAGAGG GAGGTGGAAT TCCGCGTGTA GCAGTGAAAT GCGTAGATAT GCGGAGGAAC 660
ACCGATGGCG AAGGCAGCCT CCTGGGATAA CACTGACGCT CATGCACGAA AGCGTGGGGA 720
GCAAACAGGA TTAGATACCC TGGTAGTCCA CGCCCTAAAC GATGTCAACT AGCTGTTGGG 780
GCCTTCGGGC CTTGGTAGCG CAGCTAACGC GTGAAGTTGA CCGCCTGGGG AGTACGGTCG 840
CAAGATTAAA ACTCAAAGGA ATTGACGGGG ACCCGCACAA GCGGTGGATG ATGTGGATTA 900
ATTCGATGCA ACGCGAAAAA CCTTACCTAC CCTTGACATG TCTGGAATGC CGAAGAGATT 960
TGGCAGTGCT CGCAAGAGAA CCGGAACACA GGTGCTGCAT GGCTGTCGTC AGCTCGTGTC 1020
GTGAGATGTT GGGTTAAGTC CCGCAACGAG CGCAACCCTT GTCATTAGTT GCTACGAAAG 1080
GGCACTCTAA TGAGACTGCC GGTGACAAAC CGGAGGAAGG TGGGGATGAC GTCAAGTCCT 1140
CATGGCCCTT ATGGGTAGGG CTTCACACGT CATACAATGG TCGGGACAGA GGGTCGCCAA 1200
CCCGCGAGGG GGAGCCAATC CCAGAAACCC GATCGTAGTC CGGATCGCAG TCTGCAACTC 1260
GACTGCGTGA AGTCGGAATC GCTAGTAATC GCGGATCAGC ATGTCGCGGT GAATACGTTC 1320
CCGGGTCTTG TACACACCGC CCGTCACACC ATGGGAGTGG GTTTTACCAG AAGTAGTTAG 1380
CCTAACCGCA AGGAGGGCGA TACCACG 1407
The invention provides a denitrifying achromobacter and an application idea and method thereof, and a method and a way for realizing the technical scheme are more specific, the above is only a preferred embodiment of the invention, and it should be pointed out that a plurality of improvements and modifications can be made by those skilled in the art without departing from the principle of the invention, and the improvements and modifications are also considered as the protection scope of the invention. The components not explicitly described in this embodiment can be implemented by using the prior art.
Sequence listing
<110> university of Nanjing information engineering
<120> Achromobacter denitrificans and application thereof
<160> 1
<170> SIPOSequenceListing 1.0
<210> 1
<211> 1407
<212> DNA
<213> Achromobacter denitrificans (Achromobacter denitrificans)
<400> 1
CATGCAAGTC GAACGGCAGC ACGGACTTCG GTCTGGTGGC GAGTGGCGAA CGGGTGAGTA 60
ATGTATCGGA ACGTGCCTAG TAGCGGGGGA TAACTACGCG AAAGCGTAGC TAATACCGCA 120
TACGCCCTAC GGGGGAAAGC AGGGGATCGC AAGACCTTGC ACTATTAGAG CGGCCGATAT 180
CGGATTAGCT AGTTGGTGGG GTAACGGCTC ACCAAGGCGA CGATCCGTAG CTGGTTTGAG 240
AGGACGACCA GCCACACTGG GACTGAGACA CGGCCCAGAC TCCTACGGGA GGCAGCAGTG 300
GGGAATTTTG GACAATGGGG GAAACCCTGA TCCAGCCATC CCGCGTGTGC GATGAAGGCC 360
TTCGGGTTGT AAAGCACTTT TGGCAGGAAA GAAACGTTCC GGGGTTAATA CCCCGGGAAA 420
CTGACGGTAC CTGCAGAATA AGCACCGGGC TAACTACGTG CCAGCAGCCG CGGTAATACG 480
TAGGGTGCAA GCGTTAATCG GAATTACTGG GCGTAAAGCG TGCGCAGGCG GTTCGGAAAG 540
AAAGATGTGA AATCCCAGAG CTTAACTTTG GAACTGCATT TTTAACTACC GAGCTAGAGT 600
GTGTCAGAGG GAGGTGGAAT TCCGCGTGTA GCAGTGAAAT GCGTAGATAT GCGGAGGAAC 660
ACCGATGGCG AAGGCAGCCT CCTGGGATAA CACTGACGCT CATGCACGAA AGCGTGGGGA 720
GCAAACAGGA TTAGATACCC TGGTAGTCCA CGCCCTAAAC GATGTCAACT AGCTGTTGGG 780
GCCTTCGGGC CTTGGTAGCG CAGCTAACGC GTGAAGTTGA CCGCCTGGGG AGTACGGTCG 840
CAAGATTAAA ACTCAAAGGA ATTGACGGGG ACCCGCACAA GCGGTGGATG ATGTGGATTA 900
ATTCGATGCA ACGCGAAAAA CCTTACCTAC CCTTGACATG TCTGGAATGC CGAAGAGATT 960
TGGCAGTGCT CGCAAGAGAA CCGGAACACA GGTGCTGCAT GGCTGTCGTC AGCTCGTGTC 1020
GTGAGATGTT GGGTTAAGTC CCGCAACGAG CGCAACCCTT GTCATTAGTT GCTACGAAAG 1080
GGCACTCTAA TGAGACTGCC GGTGACAAAC CGGAGGAAGG TGGGGATGAC GTCAAGTCCT 1140
CATGGCCCTT ATGGGTAGGG CTTCACACGT CATACAATGG TCGGGACAGA GGGTCGCCAA 1200
CCCGCGAGGG GGAGCCAATC CCAGAAACCC GATCGTAGTC CGGATCGCAG TCTGCAACTC 1260
GACTGCGTGA AGTCGGAATC GCTAGTAATC GCGGATCAGC ATGTCGCGGT GAATACGTTC 1320
CCGGGTCTTG TACACACCGC CCGTCACACC ATGGGAGTGG GTTTTACCAG AAGTAGTTAG 1380
CCTAACCGCA AGGAGGGCGA TACCACG 1407
Claims (9)
1. Achromobacter denitrificans @Achromobacter denitrificans) YSQ030 and the preservation number is CGMCC NO.22850.
2. Use of the denitrifying achromobacter according to claim 1 for preparing microbial agents.
3. A microbial agent comprising the achromobacter denitrificans according to claim 1.
4. A method for preparing the microbial agent of claim 3, which is characterized in that: inoculating the denitrifying achromobacter into a fermentation culture medium with the pH of 7.2-7.5, and culturing at 26+/-2 ℃ until the viable count reaches 2 hundred million/ml or more than 2 hundred million/g.
5. The method for preparing the microbial agent according to claim 4, wherein the method comprises the following steps: the fermentation medium comprises 5.0 g/L peptone, 3.0 g/L beef extract powder, 0.3 mM sodium nitrate and 4.4 mM sodium succinate, and the solvent is water; or comprises 10.0 g/L tryptone, 5.0 g/L yeast extract powder, 10.0 g/L sodium chloride, 0.3 mM sodium nitrate and 4.4 mM sodium succinate, and the solvent is water; or comprises 10.0 g/L tryptone, 5.0 g/L maltose, 10.0 g/L sodium chloride, 0.3 mM sodium nitrate and 4.4 mM sodium succinate, and the solvent is water; or 10.0 g/L soybean peptone, 10.0 g/L sucrose, 5.0 g/L sodium chloride, 0.3. 0.3 mM sodium nitrate and 4.4. 4.4 mM sodium succinate, and water as solvent.
6. Use of the denitrifying achromobacter according to claim 1 or the microbial agent according to claim 3 in the preparation of a bio-organic fertilizer.
7. The use according to claim 6, characterized in that: the biological organic fertilizer is prepared by inoculating a microbial agent to the organic fertilizer.
8. The use according to claim 7, characterized in that: the inoculation proportion of the microbial agent is 1 milliliter of microbial agent inoculated per gram of organic fertilizer.
9. Use of the achromobacter denitrificans according to claim 1 for reducing nitrous oxide emissions from farmland greenhouse gases.
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