Method for sifting functional fungoid having specific affinity with non-leguminous plant by using agglutinin

A technology of legumes and lectins, applied in biochemical equipment and methods, microbial determination/inspection, biological testing, etc., can solve problems such as lack of

Inactive Publication Date: 2007-08-08
ANHUI AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there is no information to prove that the bacteria and plants used in the production of biological fertilizers have specificity or a...

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] Example 1: Screening of cotton rhizosphere-specific functional strains

[0021] 1 Extraction of cottonseed lectin

[0022] Cottonseed contains more lectins, but the seeds also contain more fat, so it is necessary to take measures to remove the fat in the material and then extract the lectins.

[0023] ① Peel 100g cottonseed into powder;

[0024] ②Add 500ml of anhydrous ether to degreasing for 10min, repeat twice, vacuum pump to drain excess ether;

[0025] ③Add 500ml PBS (0.01mol, pH7.2) for leaching at 4°C (the leaching solution has the strongest activity at 72h);

[0026] ④ Filter to remove coarse slag, and centrifuge the filtrate (4000rpm, 20min, normal temperature) to remove fine slag;

[0027] ⑤Put the supernatant in an ice bath, add (NH 4 ) 2 SO 4 to 40% to 90% saturation (add and stir to prevent protein denaturation);

[0028] ⑥ Let stand overnight, centrifuge (6000rpm, 30min, 4℃), discard the supernatant;

[0029] ⑦ Dissolve the precipitate with 40ml of ...

Embodiment 2

[0075] Embodiment 2: Screening of wheat rhizosphere-specific potassium-decomposing strains

[0076] 1 Extraction and purification of wheat germ agglutinin

[0077] A method for the extraction of lectins from materials containing a small amount of fat is used.

[0078] ① Wheat Germ 100g

[0079] ②Add 1000ml 0.05mol·L -1 Extract with HCl, stir for 1h, at room temperature, centrifuge at 2000×g for 10min, discard the residue, and the supernatant is the crude lectin extract;

[0080] ③Put the supernatant in an ice bath, add ammonium sulfate to 40%-90% saturation, stir for 1 hour; centrifuge at 10,000g at 4°C for 15 minutes, discard the supernatant;

[0081] ④Add 45ml 0.05mol·L to the precipitate -1 Dissolve in HCl, add 15ml of n-butanol dropwise, stir for 1h to degrease, centrifuge at 3000g for 30min, keep the water phase, (repeat the degreasing twice);

[0082] ⑤ Dilute the centrifuged supernatant to 0.05mol·L -1 HCl dialyzed overnight;

[0083] ⑥Add ammonium sulfate to th...

Embodiment 3

[0103] Example 3: Screening of rice rhizosphere-specific phosphorus-solubilizing strains

[0104] 1 Extraction and purification of rice lectins

[0105] The rice lectin was extracted and purified using the same "method of extraction of lectin in a small amount of fat-containing material" as in Example 2.

[0106] 2 Labeling Rice Lectin with Biotin

[0107] ① Dialyze the purified lectin (10ml) in labeling buffer (0.1Mol / LNaAc, pH 5.5, 0.1mol / LNaCl) at 4°C overnight;

[0108] ② Pipet 5ml into a centrifuge tube, add sodium periodate solution to a final concentration of 10mmol / L, and place it on an ice bath for 30min to avoid light to oxidize the sugar groups on the lectin molecule;

[0109] ③ Equilibrate the Sephadex G25 PD-10 prepacked column with PBS, pass the oxidized lectin through the column, and collect the lectin fraction with coagulation activity;

[0110] ④ Add biotin to the lectin tube to a final concentration of 5 mmol / L, and place it on a shaker for 1 hour at 25°C;...

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PUM

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Abstract

The invention relates to the method which uses lectin to screen the function bacterium with specific affinity to the non-legume-plant. The invention uses the fluorescent tracer to label the lectin, and considers the tracer-labeled lectin as mutual recognition media between the plant and microorganism, and from the plant surface, the root surface, and the root rhizosphere, screens excellent specific bacterium trunk with high efficient nitrogen fixation, phosphor decomposition, potassium decomposition, growth promoting, and as well as promoting the growth of other plants, to be the bacterium trunk of bio-fertilizer production, to produce high efficient and specific biological fertilizer applied for the different plants, in favor of the roots plant of the bacterium trunk in a particular plant. These microorganisms can be made of plant growth regulators, soil conditioner, and so on, thus reducing the fertilizer usage in the agricultural production, and solving the problem of environmental pollution by excessive usage of fertilizer, with a view to obtaining good economic, ecological and social benefits.

Description

technical field [0001] The present invention relates to a screening method for functional bacteria with specific affinity of non-leguminous plants. Background technique [0002] Lectins are a class of glycoproteins that are widely present in various plants and have the property of agglutinating red blood cells. Lectins can recognize specific compound determinants on the surface of bacterial cells, and a lectin has the ability to specifically bind to a specific glycosyl; lectins secreted by plant roots have the ability to specifically recognize rhizosphere microorganisms. Important role; lectin has multivalent binding ability and can bind to tracers such as fluorescein and biotin. Therefore, tracers such as fluorescein can be used to label the lectin, and the labeled lectin can be used as a medium for mutual recognition between plants and microorganisms, and the microorganisms that can be recognized by the lectin can be displayed under a microscope or other technical methods...

Claims

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Application Information

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IPC IPC(8): G01N33/58G01N33/53G01N21/64C12Q1/02
Inventor 唐欣昀
Owner ANHUI AGRICULTURAL UNIVERSITY
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