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Bacillus subtilis for controlling citrus canker

A Bacillus subtilis, canker technology, applied in the direction of chemicals for biological control, microorganism-based methods, bacteria, etc., can solve problems such as harming ecological safety and human health, enhancing drug resistance of pathogenic bacteria, and environmental pollution. Good control effect

Inactive Publication Date: 2013-06-26
SOUTHWEST UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] At present, the control of citrus canker is still dominated by chemical agents, which leads to serious consequences such as increased resistance of pathogenic bacteria, environmental pollution, etc., and finally poor control effect, Hazards to ecological security and human health

Method used

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  • Bacillus subtilis for controlling citrus canker
  • Bacillus subtilis for controlling citrus canker
  • Bacillus subtilis for controlling citrus canker

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Example 1. Identification of Bacillus subtilis CCCQ080 strain

[0030] (1) Morphological observation and identification: culture the strain CCCQ080 on the NA culture plate at 28°C for 1-3 days, and observe the colony morphology (shape, color luster, whether the surface is raised or sunken, edge shape, etc.);

[0031] (2) Observation of physiological and biochemical reactions: Inoculate the strain CCCQ080 in the test medium for temperature, pH, salt tolerance, sugar alcohol fermentation, citrate utilization, nitrogen compound utilization, starch and other macromolecular compound decomposition and enzyme production, etc. observe the reaction;

[0032] (3) Molecular biological identification: extract the genomic DNA of the strain, use the bacterial universal primer fD1: 5'-AGAGTTTGATCCTGGCTC AG-3', rP1: 5'-ACGGTTACCTTGTTA CGACTT-3', and use the genomic DNA of the strain CCCQ080 as a template for PCR After amplification, it was detected by electrophoresis and sequenced; th...

Embodiment 2

[0033] Embodiment 2. Bacillus subtilis CCCQ080 bacterial strain disease control effect

[0034] 2.1 Bacteriostatic effect of CCCQ080 strain: inoculate the strain on NA plates, culture at a constant temperature of 28°C for 2 days, and spray 3×10 8 The cfu / mL suspension of X. citri was sprayed on the plate (about 100 μL / dish), and repeated 3 times. After culturing at a constant temperature of 28°C for 3 days, measure the diameter of the inhibition zone. To determine the inhibitory effect of CCCQ080 strain on citrus canker.

[0035] 2.2 Disease control effect of CCCQ080 strain

[0036] Detached leaf method: take some healthy sweet orange citrus leaves, wash the surface with distilled water, and dry. Soak in 75% ethanol solution for 30 s, then wash with sterile water and dry. The leaves were randomly and evenly divided into 4 treatments of A, B, C and D. In treatment A, only sterilized water was sprayed on the leaves; in B, only the fermentation broth of antagonistic bacteria ...

Embodiment 3

[0038] Example 3. Analysis of metabolites of CCCQ080 strain

[0039] 3.1 Detection of chitinase activity

[0040] Sole carbon source medium (Chi-Ayers): 1.0g ammonium dihydrogen phosphate, 0.2g magnesium sulfate, 0.2g potassium chloride, 20g agar, 1% colloidal chitin to 1000mL, pH: 7.0.

[0041] Preparation of colloidal chitin: Dissolve 20g of chitin in 350mL of concentrated hydrochloric acid, place at 4°C for 24h, filter through glass wool, add 2L of ice-free ethanol to the filtrate overnight at -20°C, centrifuge at 10,000 r / min for 20min, rinse with tap water Precipitate to neutral pH, freeze-dry, and store in a sealed seal at -20°C. When using colloidal chitin, grind it more than 5 times with a manual homogenizer.

[0042] Inoculate the antagonistic bacteria on the plate made of the above medium, culture at 28°C for 7-10 days, observe whether there is a transparent circle and measure the size of the transparent circle, if there is a transparent circle, it indicates that t...

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Abstract

The invention relates to a bacillus subtilis CCCQ080 with stronger activity prevention for citrus canker, belonging to the field of agriculture disease biological control. The bacillus subtilis is obtained by being separated from natural environment host leaves and is identified according to the morphological characteristics and molecular biology. The bacillus subtilis has very strong inhibition effect on the citrus canker, and also has good control effect on the citrus canker. Furthermore, the bacillus subtilis also has broad-spectrum antagonistic activity, has stronger inhibition effect on pathogenic bacteria such as citrus canker pathogen and tobacco angular leaf spot pathogen, and also has good inhibition effect on pathogenic fungus such as tobacco botrytis cinerea, apple canker pathogenic bacteria and thielaviopsis basicola. The secondary metabolite of the bacillus subtilis contains cellulase, protease, siderophore and the like. The biological film of the canker pathogenic bacteria can be destroyed by the fermentation filtrate of the bacillus subtilis, and the environment-friendly biopesticide can be produced by bacterial fermentation or secondary metabolite extraction technology, so that the bacillus subtilis has important business development and application value.

Description

technical field [0001] The invention relates to the technical field of biological control of plant diseases, in particular to a bacillus subtilis CCCQ080 strain used for preventing and treating citrus canker. Background technique [0002] Citrus is the largest fruit in the world, and it is planted in 138 countries (regions) including Brazil, China, and the United States, with a total area of ​​about 8 million hectares 2 , with a total output of more than 120 million tons. China is the most important origin center of citrus, and the current citrus cultivation area is about 1.9 million hectares 2 , with an annual output of about 20 million tons, ranking first and second in the world respectively. Citrus canker is caused by Xanthomonas rugosa Xanthomonas axonopodis pv. cirri ) caused destructive diseases of citrus, which is a major quarantine object at home and abroad, mainly infecting Rutaceae Citrus, Citrus and Golden Citrus plants. The disease is mainly di...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20A01P1/00C12R1/125
Inventor 马冠华肖崇刚董国菊陈国康
Owner SOUTHWEST UNIVERSITY
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