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

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

Inactive Publication Date: 2012-01-25
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 affinity, and can be suitable for the specific conditions of different crops in different regions. At present, there is a lack of related technologies.

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. It is necessary to take measures to remove the fat in the material and then extract the lectins.

[0023] ①Dehull and grind 100g cottonseed into powder;

[0024] ②Add 500ml of anhydrous ether and degreasing for 10 minutes, repeat twice, and drain the excess ether by vacuum pump;

[0025] ③Add 500ml PBS (0.01mol, pH7.2) for 4℃ extraction (72h extract is the most active);

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

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

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

[0029] ⑦The precipitate is dissolved in 40ml of distilled water, a...

Embodiment 2

[0075] Example 2: Screening of wheat rhizosphere specific potassium-releasing strains

[0076] 1 Extraction and purification of wheat germ agglutinin

[0077] The method of extracting lectin from a small amount of fat material is adopted.

[0078] ① Wheat germ 100g

[0079] ②Add 1000ml0.05mol·L -1 HCl extraction, stirring for 1h, normal temperature, centrifugation at 2000×g for 10min, discarding the residue, the supernatant is the crude lectin extract;

[0080] ③Place the supernatant in an ice bath, add ammonium sulfate to 40%-90% saturation, stir for 1 hour; centrifuge at 10000g for 15 minutes at 4°C, discard the supernatant;

[0081] ④Add 45ml0.05mol·L to the precipitation -1 Dissolve in HCl, add 15ml n-butanol dropwise, stir for 1h to degrease, centrifuge at 3000g for 30min, retain the water phase (repeated degreasing twice);

[0082] ⑤Compare the centrifugal supernatant to 0.05mol·L -1 HCl dialysis overnight;

[0083] ⑥Add ammonium sulfate to the sample after dialysis to reach 40%-70% ...

Embodiment 3

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

[0104] 1 Extraction and purification of rice lectin

[0105] The same "method for extracting lectins from materials containing a small amount of fat" as in Example 2 was used to extract and purify rice lectins.

[0106] 2 Labeling rice lectin with biotin

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

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

[0109] ③ Equilibrate the Sephadex G25 PD-10 pre-packed column with PBS, pass the oxidized lectin through the column, and collect the lectin components with clotting activity;

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

[0111] ⑤ Equilibra...

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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 environmentalpollution by excessive usage of fertilizer, with a view to obtaining good economic, ecological and social benefits.

Description

Technical field [0001] The invention relates to a method for screening functional bacteria with specific affinity for non-leguminous plants. Background technique [0002] Lectin (Lectin) is a type of glycoprotein that is widely present in various plants and has the property of agglutinating red blood cells. Lectins can recognize specific compound determinants on the surface of bacterial cells. A lectin has the ability to specifically bind to a specific sugar group; the lectin secreted by plant roots can 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. The existing patents...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/58G01N33/53G01N21/64C12Q1/02
Inventor 唐欣昀
Owner ANHUI AGRICULTURAL UNIVERSITY
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