Botrytis cinerea antagonistic strain and screening method and application thereof

A technology of Botrytis cinerea and screening method, which is applied in the field of microbial technology and biological control, and can solve problems such as environmental pollution

Active Publication Date: 2017-03-08
BIOLOGY INST OF HEBEI ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, chemical pesticides such as pyrimethanil, azoxystrobin, carbendazim, dimethocarb, and saccharin are mainly used for prevention and control. The large-scale use of chemical pesticides has not only caused environmental pollution, but also caused botrytis cinerea to affect pyrimella.

Method used

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  • Botrytis cinerea antagonistic strain and screening method and application thereof
  • Botrytis cinerea antagonistic strain and screening method and application thereof
  • Botrytis cinerea antagonistic strain and screening method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] Example 1 Isolation, Purification and Preservation of Bacteria in Soil

[0035] (1) 120 soils collected from Yunnan, Heilongjiang, Xinjiang, Hebei and other places were used to separate the soil by dilution plate method using PB medium. After culturing in the incubator at 28°C for 3 days, the colonies with different shapes were picked. Bacteria were streaked and purified five times, numbered, and stored in a -80°C refrigerator with 15% glycerol.

[0036](2) Use the plate confrontation method for primary screening of antagonistic bacteria. Use a puncher with a diameter of 5 mm to punch holes on the edge of the cucumber gray mold, inoculate the bacteria block in the center of the PDA plate, and pick 4 different single colonies evenly on the At a position 3cm away from the indicator bacteria, after culturing in the dark at 22°C for 5 days, the antibacterial effect was observed, and the strains with the antibacterial zone were selected for re-screening. 4,200 strains of ba...

Embodiment 2

[0038] Example 2 Identification of antagonistic bacteria BA-KA4

[0039] (1) Culture characteristics and morphological characteristics of antagonistic bacteria

[0040] The strain BA-KA4, as image 3 As shown, the growth on PB medium does not produce pigment, and the colonies are flat or round, milky white and opaque, with irregular edges and wrinkles on the surface. Gram-positive, microscopically observed as rod-shaped, producing spores, spores are oval or cylindrical.

[0041] (2) Physiological and biochemical characteristics of antagonistic bacteria

[0042] Refer to "Berjamin Bacterial Identification Manual" and "Common Bacterial System Identification Manual": V-P test positive, methyl red test negative, indole test positive, gelatin liquefaction test positive, citrate utilization test positive, starch hydrolysis test positive , lipase negative, H 2 S production test positive, nitrate reduction test positive, malonate utilization negative, tyrosine hydrolysis negative, ...

Embodiment 3

[0049] Example 3 Pot control effect test of Bacillus amyloliquefaciens BA-KA4 on cucumber gray mold

[0050] To prepare a seed bottle, pick a single colony of BA-KA4 and inoculate it into a 100 mL Erlenmeyer flask filled with 20 mL of PB liquid medium, and culture it on a shaker at 28°C and 180 rpm for 24 hours. Draw 10 mL of the bacterial solution from the seed bottle and inoculate it into a 500 mL Erlenmeyer flask containing 200 mL of PB liquid medium. After 48 hours of shaking culture at 28 ° C and 180 rpm on a shaking table, dilute it with sterile water to 1 × 10 8 CFU / mL is the bacterial suspension. Take the same concentration of bacterial suspension and sterilize at 121°C for 20 minutes, which is the sterile fermentation broth. Azoxystrobin was diluted 1000 times with sterile water. Use a hole puncher to punch holes on the edge of Botrytis cinerea growing for 3 days, put the bacteria block into 0.9% normal saline, 22°C, 180rpm shaker shaker for 30min, dilute with steri...

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Abstract

The invention belongs to the fields of microbial technology and biological control and particularly relates to bacillus amyloliquefaciens BA-KA4 and a screening method and application thereof. Bacteria are separated from soil, and an agar disk diffusion method is adopted for screening BA-KA4 out of the bacteria, wherein the inhibition ratio of BA-KA4 on cucumber botrytis cinerea is 65%, and BA-KA4 has stable inheritance. Through cultural characteristics, morphology and physiological and biochemical characteristics of the strain in combination with result reanalysis of a 16S rDNA sequence and a gyrB gene sequence, it is preliminarily identified that the strain BA-KA4 is bacillus amyloliquefaciens. The strain is stored with the accession number of CGMCC No.12190 in the Common Microorganism Center of China General Microbiological Culture Collection Center. The strain has the best control effect on the cucumber botrytis cinerea, a long lasting period and a broad antibacterial spectrum.

Description

technical field [0001] The invention belongs to the field of microbial technology and biological control, and specifically relates to a Bacillus amyloliquefaciens (Bacillus amyloliquefaciens) BA-KA4 and a screening method and application thereof. Background technique [0002] Botrytis cinerea is a worldwide plant disease, which is caused by the fungus Botrytis cinerea, which infects various crops including cucumbers, tomatoes, grapes, etc. serious threat to the yield and quality of vegetables and fruits. At present, chemical pesticides such as pyrimethanil, azoxystrobin, carbendazim, dimethocarb, and saccharin are mainly used for prevention and control. The large-scale use of chemical pesticides has not only caused environmental pollution, but also caused botrytis cinerea to affect pyrimella. The resistance of amine, carbendazim, and diethofencarb to other agents, Wang Meiqin et al. have shown that the resistance of cinerea cinerea in Jinzhong area of ​​Shanxi Province to D...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12N1/02A01N63/02A01P3/00C12R1/07
CPCA01N63/10C12N1/02C12N1/205C12R2001/07
Inventor 刘昆昂黄亚丽贾振华马宏宋聪赵芊黄媛媛刘方张立文宋水山
Owner BIOLOGY INST OF HEBEI ACAD OF SCI
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