CN114561314B - Streptomyces corollineus A1 and application thereof - Google Patents
Streptomyces corollineus A1 and application thereof Download PDFInfo
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- CN114561314B CN114561314B CN202111066705.XA CN202111066705A CN114561314B CN 114561314 B CN114561314 B CN 114561314B CN 202111066705 A CN202111066705 A CN 202111066705A CN 114561314 B CN114561314 B CN 114561314B
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Classifications
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
- A01N63/20—Bacteria; Substances produced thereby or obtained therefrom
- A01N63/28—Streptomyces
-
- 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
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P3/00—Preparation of elements or inorganic compounds except carbon dioxide
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- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Zoology (AREA)
- Organic Chemistry (AREA)
- Wood Science & Technology (AREA)
- Biotechnology (AREA)
- General Health & Medical Sciences (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Genetics & Genomics (AREA)
- Microbiology (AREA)
- General Engineering & Computer Science (AREA)
- Virology (AREA)
- Biochemistry (AREA)
- Tropical Medicine & Parasitology (AREA)
- Biomedical Technology (AREA)
- Medicinal Chemistry (AREA)
- Agronomy & Crop Science (AREA)
- Pest Control & Pesticides (AREA)
- Plant Pathology (AREA)
- Dentistry (AREA)
- Environmental Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The application relates to the technical field of microorganisms, in particular to streptomyces corset A1 and application thereof. The preservation number of the streptomyces coronensis (Streptomyces bottropensis) A1 is CCTCC NO. M2021659. The streptomyces corollosus A1 can reduce selenite in a culture medium into elemental selenium, the streptomyces corollosus A1 can synthesize nano-selenium, fat, protein and other components, and the existence of the substances can enhance the stability and biological activity of the synthesized nano-selenium, improve the germination rate of seeds and promote plant growth.
Description
Technical Field
The application relates to the technical field of microorganisms, in particular to streptomyces corset A1 and application thereof.
Background
Actinomycetes (Actinomycetes) are a group of gram-positive, high (g+c) molar content (> 55%) bacteria, a special group of prokaryotes that can form branching hyphae and conidia. Actinomycetes are widely distributed in nature, exist in soil, air and water mainly in spore or mycelium states, and especially have low water content, rich organic matters and most amount in neutral or slightly alkaline soil. Actinomycetes grow in hyphae, mainly propagate as spores, and are named because colonies are radial. Most actinomycetes have developed branch hyphae, which are fine and have a width close to that of the rod-shaped bacteria, and are about 0.2 to 1.2 micrometers. The method can be divided into: the main function of the nutritional hyphae, also called endo hyphae or primary hyphae, is to absorb nutrient substances, and some of the nutritional hyphae can produce different pigments, which is an important basis for strain identification; aerial hyphae are stacked on nutritional hyphae, which are also called secondary hyphae; spore filaments, aerial hyphae develop to a certain stage, on which spore-forming hyphae can differentiate. The colony of actinomycetes consisted of mycelium. Generally circular, smooth or with many folds, the colony is radial.
Selenium is a trace element necessary for human body, has special physiological function and wide pharmacological action, and if selenium is deficient, diseases such as cardiovascular diseases, liver diseases, cancers, and bone joint diseases are easily caused, the intake of selenium is lower than the physiological requirement, and proper amount of selenium is needed to be supplemented, so that the immunity of the organism is enhanced. Most of the existing means for supplementing selenium to vegetables are to spray leaves with sodium selenite or apply selenium mineral powder to root systems, wherein the selenite in the selenium source has high toxicity, and further development of selenium supplementing products is restricted.
Compared with a physical and chemical synthesis method, the method for synthesizing the nano-selenium by utilizing the microorganism is an environment-friendly synthesis way, and has the advantages of mild conversion condition, high safety, environmental protection, economy, more stable structure, good dispersibility and the like. At present, microorganisms for synthesizing nano-selenium are mainly concentrated on bacteria and fungi, and only streptomyces tanggdeiensis (Streptomyces tendae) is found in actinomycetes to have the capability of synthesizing nano-selenium.
Disclosure of Invention
The application provides a Streptomyces corchocola A1 and application thereof, and metabolites of the Streptomyces corchocola A1 can improve seed germination rate and promote plant growth.
In a first aspect, the present application provides a strain of streptomyces corset (Streptomyces bottropensis) A1, wherein the collection number of the streptomyces corset A1 is cctccc No. m2021659.
Optionally, the gene sequence of the streptomyces coronensis A1 is shown in SEQ ID NO: 1.
Optionally, the metabolite of the streptomyces coronensis A1 comprises nano-selenium.
In a second aspect, the present application provides a microbial agent comprising streptomyces coronensis (Streptomyces bottropensis) A1, the microbial agent comprising: streptomyces corollosus A1 with the preservation number of CCTCC No. M2021659 or/and a culture or/and a processed product thereof.
Optionally, the microbial agent is obtained by inoculating the streptomyces corollosus A1 into a culture medium for culture, and the selenium source in the culture medium comprises sodium selenite.
The use of the streptomyces corollosus A1 according to the first aspect, or the microbial agent according to the second aspect, for the production of elemental selenium in a selenium-containing medium.
The use of the microbial agent of the first aspect of the streptomyces corollosus A1 or the second aspect of the microbial agent for seed germination or plant growth or inhibition of soft rot pathogen of konjak.
Optionally, the plant growth comprises: the plant height is increased, and/or the length of main roots is increased, and/or the number of roots is increased, and/or the fresh weight of seedlings is increased.
In a third aspect, the application provides a treatment method for reducing selenium, which comprises the steps of inoculating the streptomyces corset A1 in the first aspect or the microbial agent in the second aspect into a solution containing sodium selenite for selenium reduction.
Compared with the prior art, the technical scheme provided by the embodiment of the application has the following advantages:
according to the streptomyces corollosus (Streptomyces bottropensis) A1, the preservation number of the streptomyces corollosus A1 is CCTCC (China center of China) No. M2021659, selenite in a culture medium can be reduced into elemental selenium by the streptomyces corollosus A1, nano selenium (components such as selenium, fat and proteins) can be synthesized by the streptomyces corollosus A1, stability and biological activity of the synthesized nano selenium are enhanced due to the existence of the substances, germination rate of seeds can be improved, plant growth is promoted, and pectobacterium chrysanthemi in the konjak soft rot pathogen can be inhibited.
Drawings
The accompanying drawings, which are incorporated in and constitute a part of this specification, illustrate embodiments consistent with the invention and together with the description, serve to explain the principles of the invention.
In order to more clearly illustrate the embodiments of the invention or the technical solutions in the prior art, the following description will briefly explain the drawings used in the embodiments or the description of the prior art, and it will be obvious to those skilled in the art that other drawings can be obtained from these drawings without inventive effort.
FIG. 1 shows colony morphology of strain A1 provided in example 1 of the present application on different media;
FIG. 2 is a phylogenetic tree constructed from Streptomyces corset A1 and Streptomyces related strains based on the 16SrRNA sequence in example 1 of the present application;
FIG. 3 shows the addition of Na at various concentrations in example 2 of the present application 2 SeO 3 Selenium-rich properties of Streptomyces coronensis A1 under conditions
FIG. 4 is a scanning electron microscope photograph of the cell morphology of strain A1 and its synthesized nano-selenium in example 3 of the present application;
FIG. 5 is an X-ray energy spectrum analysis of strain A1 and its synthesized nano-selenium in example 3 of the present application;
FIG. 6 is a graph showing analysis of elemental composition of spherical particles of nanoselenium by energy spectrometer (EDS) in example 3 of the present application;
FIG. 7 shows sodium selenite Na in example 3 of the present application 2 SeO 3 (A) X-ray diffraction pattern (XRD) with biological nano-selenium SeNPs (B);
FIG. 8 is a Fourier infrared transform spectrum of biological nano-selenium Senps in example 3 of the present application;
FIG. 9 is a graph showing the effect of different levels of selenium-enriched fermentation filtrate on wheat (A) and corn (B) seed germination and growth in example 4 of the present application;
FIG. 10 shows the inhibitory effect of selenium-enriched fermentation filtrate on konjak soft rot pathogen in example 5 of the present application.
Detailed Description
For the purposes of making the objects, technical solutions and advantages of the embodiments of the present application more clear, the technical solutions of the embodiments of the present application will be clearly and completely described below with reference to the drawings in the embodiments of the present application, and it is apparent that the described embodiments are some embodiments of the present application, but not all embodiments. All other embodiments, which can be made by one of ordinary skill in the art without undue burden from the present disclosure, are within the scope of the present application based on the embodiments herein.
Streptomyces corollosus (Streptomyces bottropensis) A1 has the accession number: cctccc No. m2021659; the classification is named: streptomyces corollarius (Streptomyces bottropensis) A1; the preservation date is: 2021, 6 and 2; the preservation units are as follows: china Center for Type Culture Collection (CCTCC); the preservation unit addresses are: chinese university of Wuhan and Wuhan.
EXAMPLE 1 isolation and characterization of Streptomyces coronensis (Streptomyces bottropensis) Strain A1
The Streptomyces corollarius (Streptomyces bottropensis) A1 strain is isolated from the root surface soil of 8 month old healthy konjak in an agronomic greenhouse of the Ankang university. Shaking off a large amount of soil attached to the root system of healthy konjak, washing a small amount of soil still attached within 2mm of the root surface into a triangular flask containing a small amount of glass beads and 90mL of sterile water, shaking on a shaker for 30min, and diluting to 10 times according to 10-fold gradient -3 ～10 -5 Respectively and uniformly coating 50 mu L of each gradient soil suspension on beef extract peptone agar medium (beef extract 3g, peptone 10g, naCl 5g, brown sugar 10g, distilled water 1000mL, agar 15g, regulating pH to 7.2-7.5 with 10% NaOH), and selecting actinomycetes of different forms for purification and preservation.
Morphological characterization of strain A1 on different media: in Gao's first culture medium (soluble starch 20.0g, naCl 0.5g, KNO) 3 1.0g，K 2 HPO 4 0.5g，MgSO 4 ·7H 2 O 0.5g，FeSO 4 0.02g, 15g of agar, 1000mL of distilled water, pH 7.2-7.4), GYM medium (yeast extract 2g, glucose 4g, malt flour 10g, caCO 3 2g, 1000mL of distilled water, pH 7.5), desert fungus culture medium (yeast powder 4g, glucose 4g, malt powder 5g, trace salt 1mL, vitamin 1mL, distilled water 1000mL, pH 7.0) and ISP2 culture medium (yeast extract 4g, malt extract 10g, glucose 4g, trace salt 1mL, distilled water 1000mL, pH 7.2) were inoculated with Streptomyces corotrichum A1, cultured at 28℃for 10d at constant temperature, and the basic of the strain A1 on the different culture mediums was observedFeatures. The basic characteristics of the Streptomyces corollosus A1 strain obtained in this test on different culture media are as follows:
the physiological and biochemical characteristic analysis results are as follows: the carbon source utilization and the physiological and biochemical characteristics of the strain A1 are matched with those of the streptomyces corollosus (Streptomyces bottropensis). Streptomyces corollosus A1 can utilize carbon sources of fructose, lactose, maltose and D-mannitol, and D-xylose, inositol and sorbitol are poor, so that gelatin can be liquefied, milk is not coagulated and peptonized, and H can be produced 2 S and melanin, the starch has strong hydrolysis capability and weak cellulose decomposition capability.
16SrDNA sequence analysis: the total DNA of the streptomyces corollosus A1 is extracted by adopting an enzymolysis method, and the bacterial 16S rDNA universal primer (PA: 5'-AGAGTTTGATCCTGGCTCAG-3'; PB:5'-AAGGAGGTG ATCCAGCCGCA-3') is utilized for PCR amplification, so as to obtain a fragment with the length of 1400-1500 bp. The amplified products were sequenced by biological engineering (Shanghai) Inc. The total length of the 16S rDNA sequence of the streptomyces corollosus A1 is 1462bp, the obtained sequence is spliced and aligned (shown as SEQ ID NO: 1), the sequence similarity search is carried out from a GenBank database by adopting a Blast method, 8 related typical strains of streptomyces with higher similarity with A1 are called, homology analysis is carried out by ClustalX v2.2 software, and a phylogenetic tree is constructed by adopting a Neighbor-Joing method in Mega 5.0 software. As can be seen from FIG. 2, the similarity of the strain A1 to Streptomyces corollosus (Streptomyces bottropensis) on the same phylogenetic branch is 98.3%, and the phylogenetic tree constructed from the Streptomyces corollosus A1 and the Streptomyces corollosus related strain based on the 16S rRNA sequence is shown in FIG. 2.
Comprehensively, according to the analysis result of the 16S rDNA sequence and the morphological, physiological and biochemical characteristics, the strain A1 is identified as the streptomyces corchowensis.
EXAMPLE 2 Synthesis of biological nanoselenium by Strain A1 in solid and liquid Medium
The strain A1 synthesizes biological nano selenium: the metabolite of the streptomyces corollosus A1 in the first aspect comprises nano-selenium, wherein the nano-selenium contains aliphatic and proteinaceous organic components. The presence of these substances enhances the stability and biological activity of the synthesized nanoselenium.
Synthesizing biological nano selenium on a solid culture medium of Gaoshan No. one: in solid medium of Gaoshi No. one (soluble starch 20.0g,NaCl 0.5g,KNO) 3 1.0g，K 2 HPO 4 0.5g，MgSO 4 ·7H 2 O 0.5g，FeSO 4 0.02g, 15g of agar, 1000mL of distilled water and pH 7.2-7.4) of Na with a certain mass 2 SeO 3 Since the microorganism can make Na 2 SeO 3 The microbial cells can deposit red particles after being reduced into red biological selenium, and the selenium enrichment capacity of the strain can be judged by the depth of red and the selenium content in the culture medium. Culture medium Na tolerated by Streptomyces corollosus A1 strain 2 SeO 3 As can be seen from FIG. 3, A-D, A. Control (no Na was added to the medium 2 SeO 3 ) The method comprises the steps of carrying out a first treatment on the surface of the B. Addition of 10mg/LNa 2 SeO 3 The method comprises the steps of carrying out a first treatment on the surface of the C. 100mg/LNa was added 2 SeO 3 The method comprises the steps of carrying out a first treatment on the surface of the D. 200mg/LNa was added 2 SeO 3 The method comprises the steps of carrying out a first treatment on the surface of the Streptomyces corollosus A1 control group and Na 2 SeO 3 The color of the group bacterial liquid is different; the mycelium of the streptomyces corollosus A1 appears white under the selenium-free culture condition; at Na (Na) 2 SeO 3 On a plate with the concentration of 10mg/L, the lawn of the streptomyces coronensis A1 is light red; at Na (Na) 2 SeO 3 The Streptomyces corollosus A1 lawn on the plates with the concentration of 100mg/L and 200mg/L shows dark red, which indicates that the Streptomyces corollosus A1 synthesizes red elemental selenium under the selenium-containing condition.
Synthesizing biological nano selenium on a liquid culture medium of Gaoshan No. 1: liquid culture medium of Gaoshi No. 1 (soluble starch 20.0g,NaCl 0.5g,KNO) 3 1.0g，K 2 HPO 4 0.5g，MgSO 4 ·7H 2 O 0.5g，FeSO 4 0.02g, distilled water 1000mL, pH 7.2-7.4) in 250mL triangular flask (100 mL/bottle), PE film sealing, sterilizing with 121 deg.C high pressure steam for 20min, adding Na with pore diameter of 0.22 μm and sterile filter membrane in ultra clean bench 2 SeO 3 Solution, na in the culture medium 2 SeO 3 The concentration reaches 200mg/L, and the control group is not added with Na 2 SeO 3 . Mycelium pellet was selected from activated strain A1 slant and inoculated to control and 200mg/L Na added, respectively 2 SeO 3 In Gao's liquid culture medium No. one, each treatment is repeated 3 times, the culture is continuously carried out for 10 days at the constant temperature of 150r/min and 28 ℃, the bacterial liquid is red in color (figure 4), and the same shows that elemental selenium is gradually generated. In order to further determine the accuracy of the result, the dry powder of the thalli is taken for scanning electron microscope observation.
Example 3 cell morphology observations of Strain A1 and characterization of nanoselenium essential characteristics
The morphology of the strain A1 cells cultured in liquid culture under Gao's first protocol was observed by Scanning Electron Microscopy (SEM). Inoculating activated strain A1 mycelium blocks to non-added Na 2 SeO 3 200mg/L Na was added 2 SeO 3 In Gao's liquid culture medium with aperture of 0.22 μm and sterile filter membrane filtration, 150r/min and 28 deg.C constant temperature continuous shaking culture are carried out for 10d, the culture is centrifugated in 50mL centrifuge tube at 4 deg.C and 10000r/min for 10min, the obtained bacterial precipitate is subjected to a series of treatments such as freeze drying, metal spraying, etc., and then the cell morphology of the strain A1 and the culture thereof is observed by a scanning electron microscope. As can be seen from the SEM photograph presented in fig. 5, the strain A1 cell morphology mag=10.0 KX in fig. 5A; in figure 5B, strain A1 synthesizes nano selenium form mag=10.0 KX; strain A1 cell morphology mag=20.0 KX in panel 5C; in the 5D graph, the strain A1 synthesizes nano selenium form mag=20.0 KX; from the 5B and 5D graphs, it can be seen that Na was added 2 SeO 3 The treatment group of (2) produces the simple substance spherical granular biological nano selenium.
The analysis of the elemental composition of the spherical nano-selenium particles surface by using an energy spectrometer (EDS) shows the result that the left graph is the X-ray energy spectrum analysis of the strain A1, the right graph is the X-ray energy spectrum analysis of the nano-selenium synthesized by the strain A1, the characteristic peak of Se element appears in the right graph at 1.4keV, the relative content reaches 15.0%, and in addition, the spherical nano-selenium particles generated by the strain A1 are more determined to be biological nano-selenium by containing part of organic elements from biological macromolecules.
Separation of nano-selenium crystal form and surface functional group structure biosynthesized by strain A1 by X-ray diffractometer (XRD) and Fourier infrared transformation spectrum (FTIR)And (5) separating. Inoculating activated strain A1 mycelium block into 200mg/L sterile filter membrane (pore size 0.22 μm) containing filtered Na 2 SeO 3 In the liquid culture medium of Gao's No. one, 150r/min, continuous shaking culture at constant temperature of 28 ℃ for 10d, centrifugation at 4 ℃ and 10000r/min for 10min, collecting precipitate, cleaning for 3 times, refrigerating at-80 ℃ for 16h, and performing XRD and FTIR analysis respectively after vacuum freeze drying for 16 h.
From Na 2 SeO 3 As can be seen from the diffraction pattern 7A of (2) a plurality of sharp peaks appear at positions of about 22.01 DEG and 37.64 DEG of Two-Theta, but after biotransformation by the strain A1 of Streptomyces corzedoariae, a plurality of sharp peaks completely disappear, a wider peak appears at three positions of 12.91 DEG, 19.62 DEG and 29.00 DEG, no obvious crystal diffraction peak appears, which indicates a substrate Na 2 SeO 3 The crystal structure of (a) is converted into amorphous bionano-selenium by the action of the strain A1 of streptomyces coronensis (see figure 7).
Detecting whether the biological nano selenium synthesized by the strain A1 has biological macromolecules such as proteins on the surface, and analyzing the surface functional group structure by adopting Fourier infrared transformation spectrum. As can be seen from FIG. 8, at 3259.89cm -1 An absorption peak is-NH or-OH stretching vibration, and is derived from cellulose or polysaccharide substances; at 2914.51cm -1 The absorption peak is C-H stretching vibration, which is derived from fat substances; at 1638.71cm -1 And 1547.08cm -1 The absorption peaks of the (C) are respectively corresponding to the stretching vibration of-CONH-and N-H and are derived from protein substances; at 1406.10cm -1 The absorption peak at the position belongs to aliphatic C-H stretching vibration; at 1029.00cm -1 The absorption peak at the position corresponds to the C-O stretching vibration, which shows that the strain A1 converts Na according to the analysis result of the Fourier infrared transformation spectrum according to the spectroscopy and the spectrum analysis of polysaccharide substances 2 SeO 3 In the process, the secreted polysaccharide substance provides binding sites for the formation of nano-selenium particles. In addition, the strain A1 synthesizes nano-selenium and contains aliphatic and proteinaceous organic components, and the existence of the substances enhances the stability and the biological activity of the synthesized biological nano-selenium.
Example 4 detection of the Effect of Strain A1 selenium-enriched fermentation filtrate on seed germination and the Effect on plants
Preparing selenium-enriched fermentation filtrate of the strain A1: inoculating activated strain A1 mycelium block into 200mg/L Na 2 SeO 3 Continuously shaking and culturing the solution (with a sterile filter membrane with a pore size of 0.22 μm) in Gao's first liquid culture medium at constant temperature of 150r/min and 28 ℃ for 10d, centrifuging the culture in a 50mL centrifuge tube at 4 ℃ and 10000r/min for 10min, and collecting the supernatant selenium-enriched fermentation filtrate.
Life-promoting detection of selenium-rich fermentation filtrate of strain A1: since the sexual propagation organ of konjak is a solid seed, but the dormancy period is long, the emergence rate is low, and the microorganism metabolite life-promoting test is difficult to directly use the solid seed and seedling. Wheat and corn seeds and seedlings are sensitive to the reaction of active substances in microbial metabolites and are easy to measure under laboratory conditions, and the growth-promoting activity measured by the wheat and corn seeds and seedlings has good reproducibility on crops. Therefore, the indoor culture dish bioassay is adopted to detect the effect of the selenium-enriched fermentation filtrate of the strain A1 on the germination of wheat and corn seeds and the growth of plants. Sterilizing the surface of the tested wheat and corn seeds with 70% alcohol for 30s, washing with sterile water for 3 times, sterilizing with 10% (v/v) NaOCl surface for 1min, washing with sterile water for 3 times, placing into test tubes, and respectively using sterile water (control) for 10 grains in each test tube; 200mg/L Na was added 2 SeO 3 The strain A1 of (2) was immersed in the cell-free fermentation filtrate 0 (stock solution), 5-fold and 50-fold dilutions at 25℃for 24 hours in the dark. Discarding the immersion liquid, washing the seeds with sterile water, putting the seeds into a culture dish filled with sterile water-soaked filter paper, repeating 3 dishes for each treatment, culturing 10 seeds in each dish at constant temperature of 25 ℃ for 2 days, counting the germination rate of the seeds, culturing for 7 days, measuring the height of seedlings, the length of main roots, and counting the number of single plants and the total fresh weight.
As can be seen from table 1 and fig. 9A, the selenium-enriched fermentation filtrate stock solution treatment of strain A1 significantly increased the germination rate, seedling height, root length and total fresh weight of wheat seeds by 36.4%, 76.9%, 473.1% and 57.1%, respectively, compared with the control, the root length and total fresh weight of corn seedlings by 83.0% and 18.3%, respectively (P < 0.05).
Table 1 effects of different levels of selenium-rich fermentation filtrate on germination and growth of wheat and corn seeds.
Note that: the different lower case letters of the same column indicate significant differences (P < 0.05).
Data analysis of variance was performed using SPSS17.0, data expressed as "mean.+ -. Standard deviation", and difference significance test (P < 0.05) was performed using Duncan's method.
Example 5 determination of bacteriostatic Activity of selenium-enriched fermentation filtrate of Strain A1
Test target bacteria source: the konjak soft rot pathogen is pectobacterium chrysanthemi (Pectobacterium chrysanthemi) CZS-B6 strain provided by China center for type culture collection, and the preservation number is: cctccc AB 2015201.
Preparing and coating a test target bacteria suspension: the activated konjak soft rot pathogen is inoculated into beef extract peptone liquid culture medium (beef extract 3g, peptone 10g, naCl 5g, brown sugar 10g, distilled water 1000mL, pH value is regulated to 7.2-7.5 by 10% NaOH), after culturing for 18-24 h at 28 ℃, centrifugating for 10min at 4 ℃ and 12000r/min, removing supernatant, and adding sterile water to suspend bacteria. Preparation of Density 10 by absorbance A600 densitometric method 8 CFU/mL of the target bacteria suspension. And (3) inoculating 50uL of bacterial suspension to a beef extract peptone agar medium, and uniformly smearing.
Filter paper diffusion method: adding 200mg/L Na 2 SeO 3 50mL of selenium-enriched fermentation filtrate of the strain A1 is added into a sterile filter membrane with the aperture of 0.22 mu m for filtering, sterilized filter paper sheets (d=6mm) are put into the sterile filtrate after filtering for soaking for 10min, then the sterile forceps are used for picking out and putting the filter paper sheets into beef extract peptone agar medium inoculated with target bacteria, 4 filter paper sheets soaked by the sterile filtrate are put at equal intervals around each dish, and 1 filter paper sheet soaked by sterile water is put in the middle of each dish for comparison.
Oxford cup method: placing 3 oxford cups on beef extract peptone agar medium inoculated with target bacteria, taking prepared 200mg/L Na 2 SeO 3 (filtration through sterile filtration membrane having pore size of 0.22 μm) Strain A1 selenium-enriched fermentation filtrate was fed into each of 2 oxford cups with 100. Mu.L of the filtrateThe filtrate, another oxford cup, was filled with 100 μl of sterile water as a control.
The above treatments were repeated 6 times, and the diameter of the inhibition zone was observed by incubation at 28℃for 24 hours. From fig. 10, it can be seen that the bacterial fermentation filtrate of the biological nano selenium synthesized by the streptomyces corollarius strain A1 has obvious antibacterial effect on konjak soft rot pathogen. The average inhibition zone diameters of the filter paper diffusion method (A) and the oxford cup method (B) are respectively 14.6mm and 13.1mm.
It should be noted that in this document, relational terms such as "first" and "second" and the like are used solely to distinguish one entity or action from another entity or action without necessarily requiring or implying any actual such relationship or order between such entities or actions. Moreover, the terms "comprises," "comprising," or any other variation thereof, are intended to cover a non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements does not include only those elements but may include other elements not expressly listed or inherent to such process, method, article, or apparatus. Without further limitation, an element defined by the phrase "comprising one … …" does not exclude the presence of other like elements in a process, method, article, or apparatus that comprises the element.
The foregoing is only a specific embodiment of the invention to enable those skilled in the art to understand or practice the invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the generic principles defined herein may be applied to other embodiments without departing from the spirit or scope of the invention. Thus, the present invention is not intended to be limited to the embodiments shown herein but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.
Sequence listing
<110> Ankang college
<120> a strain of Streptomyces corzereus A1 and application thereof
<130> 2021-9-10
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tggcgaacgg gtgagtaaca cgtgggcaat ctgcccttca ctctgggaca agccctggaa 120
acggggtcta ataccggata caacmctykc rggcatccgw tgatggtgga aagctccggc 180
ggtgaaggat gagcccgcgg cctatcagct tgttggtgag gtaacggctc accaaggcga 240
cgacgggtag ccggcctgag agggcgaccg gccacactgg gactgagaca cggcccagac 300
tcctacggga ggcagcagtg gggaatattg cacaatgggc gaaagcctga tgcagcgacg 360
ccgcgtgagg gatgacggcc ttcgggttgt aaacctcttt cagcagggaa gaagcgaaag 420
tgacggtacc tgcagaagaa gcgccggcta actacgtgcc agcagccgcg gtaatacgta 480
gggcgcgagc gttgtccgga attattgggc gtaaagagct cgtaggcggt ctgtcgcgtc 540
ggatgtgaaa gcccggggct taaccccggg tctgcattcg atacgggcag actagagtgt 600
ggtaggggag atcggaattc ctggtgtagc ggtgaaatgc gcagatatca ggaggaacac 660
cggtggcgaa ggcggatctc tgggccatta ctgacgctga ggagcgaaag cgtggggagc 720
gaacaggatt agataccctg gtagtccacg ccgtaaacgg tgggaactag gtgttggcga 780
cattccacgt cgtcggtgcc gcagctaacg cattaagttc cccgcctggg gagtacggcc 840
gcaargctaa aactcaaagg aattgacggg ggcccgcaca agcagcggag catgtggctt 900
aattcgacgc aacgcgaaga accttaccaa ggcttgacat acaccggaaa cggccagaga 960
tggtcgcccc cttgtggtcg gtgtacaggt ggtgcatggc tgtcgtcagc tcgtgtcgtg 1020
agatgttggg ttaagtcccg caacgagcgc aacccttgtt ctgtgttgcc agcatgccct 1080
tccggggtga tggggactca caggaractg ccggggtyaa ctcggaagga aa 1132
Claims (9)
1. The Streptomyces corollosus (Streptomyces bottropensis) A1 is characterized in that the preservation number of the Streptomyces corollosus A1 is CCTCC No. M2021659.
2. The streptomyces coronensis (Streptomyces bottropensis) A1 according to claim 1, wherein the gene sequence of the streptomyces coronensis A1 is set forth in SEQ ID NO: 1.
3. The streptomyces coronensis (Streptomyces bottropensis) A1 according to claim 1, wherein the metabolite of streptomyces coronensis A1 is nano-selenium.
4. A microbial agent comprising streptomyces corollosus (Streptomyces bottropensis) A1, the microbial agent comprising: streptomyces corollosus A1 with the preservation number of CCTCC No. M2021659.
5. The microbial agent according to claim 4, wherein the microbial agent is obtained by inoculating the streptomyces corollosus A1 into a culture medium for culture, and the selenium source in the culture medium is sodium selenite.
6. Use of the microbial agent of streptomyces corset A1 according to any one of claims 1 to 3, or of claim 4 or 5, for the production of elemental selenium in selenium-containing media.
7. Use of the microbial agent of streptomyces corollosus A1 according to any one of claims 1 to 3, or of the microbial agent of claim 4 or 5 for seed germination or plant growth, or for inhibiting soft rot pathogen of konjak.
8. The use according to claim 7, wherein the plant is grown to increase plant height, increase main root length, increase root number or increase fresh weight of seedlings.
9. A method for treating reduced selenium, comprising the step of introducing the microbial agent of any one of claims 1 to 3 into a solution containing sodium selenite to reduce selenium.
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| CN103952363A (en) * | 2014-05-16 | 2014-07-30 | 华中农业大学 | Bacillus ES2-45 capable of reducing selenite to produce nano-selenium and application thereof |
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