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Inhibition of plant pathogenic bacteria gacs/gaca two-component regulatory system biocontrol strain gacai and its application

A technology of plant pathogenic bacteria and biocontrol strains, applied in the direction of plant growth regulators, bacteria, applications, etc., can solve the problems of increased drug resistance of pathogenic bacteria

Active Publication Date: 2020-08-11
GUANGXI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The mechanism of action of traditional antibiotics is generally to directly kill pathogenic bacteria, and a large amount of use will also lead to environmental safety problems such as increased resistance of pathogenic bacteria

Method used

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  • Inhibition of plant pathogenic bacteria gacs/gaca two-component regulatory system biocontrol strain gacai and its application
  • Inhibition of plant pathogenic bacteria gacs/gaca two-component regulatory system biocontrol strain gacai and its application
  • Inhibition of plant pathogenic bacteria gacs/gaca two-component regulatory system biocontrol strain gacai and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026]GacAI secondary metabolite of Bacillus subtilis inhibits expression of small RNA-encoding gene rsmB in plant pathogen Pectobacterium carotovorum subsp. carotovorum (Pcc) Z3-3.

[0027] The GacS / GacA system positively regulates the expression of the small RNA-encoding gene rsmB. The promoter region of the rsmB gene was amplified by PCR and cloned into the promoter activity detection plasmid pRG970Gm to obtain the transcriptional reporter structure of rsmB (rsmB-lacZ). The expression of the reporter plasmid was controlled by the GacS / GacA system. After culturing Bacillus subtilis GacAI at 28°C for 1 week, the supernatant was extracted with an equal volume of ethyl acetate, and the extract was rotary evaporated and resuspended in 0.1 mL of dimethyl sulfoxide (DMSO). Take 5 μL of the extract and add it to 2 mL of Pcc Z3-3(p970Gm-rsmBp) and its gacA gene mutant (p970Gm-rsmBp) (OD600=0.1). After culturing for 12 hours, detect the activity of β-galactosidase in the reporter bac...

Embodiment 2

[0030] GacAI secondary metabolites from Bacillus subtilis inhibit quorum-sensing signaling molecules in plant pathogen Pcc Z3-3.

[0031] Strain Z3-3 was cultured overnight in LB medium, then diluted 1:1000 and inoculated into LB medium containing different concentrations of GacAI secondary metabolites. After culturing at 28°C for 12 hours, the supernatant was extracted with an equal volume of ethyl acetate, and the extract was spun to dryness and redissolved in 100 μL of methanol. Take 3 μL of the sample and drop it on the detection plate of quorum sensing reporter bacteria Agrobacterium tumefaciens NTL4 (pZLR4) and X-gal, after culturing overnight, record the experimental results.

[0032] see figure 2 , using the quorum system reporter strain A. tumefaciens NTL4 (pZLR4) to detect the signal molecules of the Z3-3 quorum sensing system, it was found that adding 10 μL of GacAI extract with a concentration of 30 μg / mL to the detection plate can significantly reduce the signal...

Embodiment 3

[0034] GacAI secondary metabolites of Bacillus subtilis inhibit pathogenicity of plant pathogenic bacteria Pcc Z3-3.

[0035] In order to detect the effect of GacAI secondary metabolites on the pathogenicity of Pcc Z3-3, strain Z3-3 was cultured overnight, then transferred to fresh LB medium at a ratio of 1:1000, and cultured at 28°C until OD600=1.0. Take 5 μL of the bacterial solution and mix it with different concentrations of GacAI secondary metabolites, inoculate it on Chinese cabbage, and then measure the size of the lesion after 24 hours of moist cultivation at 28°C.

[0036] see image 3 , compared with the control, the wild strain can significantly cause Chinese cabbage leaf rot; after inoculating Chinese cabbage with 10 μL of GacAI extract with a concentration of 30 μg / mL and the wild-type strain Z3-3, the pathogenicity of Z3-3 can be significantly reduced , while 40 μL of GacAI extract with a concentration of 30 μg / mL was mixed with wild-type strain Z3-3 to inoculat...

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Abstract

The present invention provides a biocontrol strain GacAI and its application for inhibiting plant pathogenic bacteria GacS / GacA two-component control system. (CGMCC for short), deposit date: June 11, 2018, deposit number: CGMCC No.15939, the secondary metabolite of Bacillus subtilis GacAI can effectively inhibit the expression of GacS / GacA system in plant pathogenic bacteria, thereby affecting the pathogenic gene If the coding genes such as glycolic acid lyase are expressed, the suspending agent prepared from the bacillus subtilis GacAI as an active ingredient can effectively prevent and treat Chinese cabbage soft rot, and the control effect on Chinese cabbage soft rot reaches 84.1%.

Description

Technical field: [0001] The invention relates to the field of biotechnology, in particular to a biocontrol bacterial strain GacAI that inhibits the bacterial GacS / GacA two-component regulation system and its application. [0002] technical background: [0003] The ability of bacteria to survive in a specific habitat requires the coordinated expression of appropriate genes in response to environmental changes. In response to the complex external environment, bacteria have evolved a variety of sensory regulatory systems such as the GacS / GacA two-component regulatory system (TCS), which monitors changes in the external environment and makes appropriate adaptations to environmental changes. reaction. The GacS / GacA system consists of a sensor kinase (sensor kinase) GacS and a corresponding response regulator (response regulator) GacA. By interacting with signaling molecules, the sensory kinase GacS is autophosphorylated. Phosphorylated GacS transfers the phosphate group to the ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/20A01N63/22A01N25/04A01P1/00C12R1/125
CPCA01N25/04A01N63/00C12N1/205C12R2001/125
Inventor 吴小刚张博张阳梁菲
Owner GUANGXI UNIV
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