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Bacillus subtilis for inhibiting phomopsis and application thereof

A technology of Bacillus subtilis and Phomopsis, applied in the field of microorganisms, can solve the problems of few reports of microbial control agents and the like, and achieve the effect of good cellulose degradation ability

Active Publication Date: 2019-06-25
INST OF MICROBIOLOGY JIANGXI ACADEMY OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Sun Yanfang and others used 15 kinds of chemical pesticides to carry out indoor screening experiments on asparagus stem blight (Sun Yanfang et al. Determination of the toxicity of 15 kinds of pesticides to the pathogen of asparagus stem blight. 2013,33(4):71-75), Wu Wenneng tested The indoor toxicity of more than 20 kinds of fungicides to the main pathogenic bacteria of kiwifruit (Wu Wenneng et al. Isolation and identification of kiwifruit soft rot pathogens and screening of antibacterial agents. Acta Plant Protection. 2017.44(5):826-832), but microbial control agents Less reported

Method used

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  • Bacillus subtilis for inhibiting phomopsis and application thereof
  • Bacillus subtilis for inhibiting phomopsis and application thereof
  • Bacillus subtilis for inhibiting phomopsis and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] Example 1 Screening of cellulase-producing strains

[0026] Screening medium: CMC-Na 10g / L, (NH 4 ) 2 SO 4 1.4g / L, MgSO 4 0.3g / L, KH 2 PO 4 2g / L, MnSO 4 1.6mg / L, FeSO 4 5mg / L, ZnSO 4 2.5mg / L, CoCl 2 2.0mg / L, agar 20g / L, pH 7.0

[0027] Seed medium: CMC-Na 10g / L, peptone 3g / L, KH 2 PO 4 4g / L, MgSO 4 ·7H 2 O 0.03g / L, pH6.0.

[0028] Fermentation enzyme production medium: CMC-Na 10g / L, (NH 4 ) 2 SO 4 4.0g / L, MgSO 4 ·7H 2 O 0.5g / L, K 2 HPO 4 2g / L, beef extract 5g / L, peptone 10g / L (natural pH)

[0029] Preliminary screening: Weigh 10g of soil sample from the bottom of Qingshan Lake in Nanchang, Jiangxi, put it into a triangular flask filled with 90mL of sterile water, place it on a shaker at 150rpm for 30min, take it out, and dilute it step by step to 10 -1 , 10 -2 , 10 -3 , 10 -4 , 10 -5 , 10 -6 6 kinds of concentrations, the dilutions of 6 kinds of concentrations were spread on the screening medium, repeated 3 times, and cultured at 25°C f...

Embodiment 2

[0032] Example 2 Bacillus subtilis T8 degrading enzyme activity assay

[0033] The filter paper enzyme activity, endo-glucosidase activity, exo-glucosidase activity and β-glucosidase activity were determined by 3,5-dinitrosalicylic acid colorimetric method (DNS method).

[0034] 2.1 Preparation of glucose standard curve

[0035] Glucose standard solution: weigh 1 g of anhydrous glucose dried at 103°C to constant weight, and dilute to 100 mL with water;

[0036] Phosphate buffer: 0.1mol / L pH 6.0 (suitable for neutral cellulase) Weigh 121.0 g of sodium dihydrogen phosphate monohydrate and 21.89 g of disodium hydrogen phosphate dihydrate, and dissolve them in 1 L of deionized water. Adjust the pH of the solution to 6.0;

[0037] DNS reagent: Weigh 10g of 3,5-dinitrosalicylic acid, put it in about 600mL of water, gradually add 10g of sodium hydroxide, stir and dissolve in a 50°C water bath (magnetic force), then add 200g of potassium sodium tartrate, After 2 g of phenol (redist...

Embodiment 3

[0072] Embodiment 3 Bacillus subtilis T8 antibacterial experiment

[0073] Fermentation enzyme production medium: CMC-Na 10g, (NH 4 ) 2 SO 4 4.0g, MgSO 4 ·7H 2 O 0.5g, K 2 HPO 4 2g, beef extract 5g, peptone 10g (pH is naturally adjusted to 1L)

[0074] LB liquid medium: tryptone (Tryptone) 10g / L, yeast extract (Yeast-extract) 5g / L, sodium chloride (NaCl) 10g / L, adjusted to pH 7.0.

[0075] LB solid medium: tryptone (Tryptone) 10g / L, yeast extract (Yeast-extract) 5g / L, sodium chloride (NaCl) 10g / L, agar 20g / L adjusted to pH 7.0.

[0076] PDA medium: potato flour 6.0g / L, glucose 20.0g / L, agar 20.0g / L, pH 5.4-5.8

[0077] 3.1 Phomopsis

[0078] Phomopsis was spread on PDA plate, cultured at 37°C for 7 days, and the spores were washed with physiological saline, Phomopsis was the suspension of Phomopsis spores.

[0079] Inoculate the Bacillus subtilis T8 bacterial liquid into a test tube containing 5 mL fermentation enzyme production medium according to the inoculum amo...

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Abstract

The invention provides a bacillus subtilis for inhibiting phomopsis which is bacillus subtilis T8 deposited in China Center for Type Culture Collection on December 5, 2018 with the preservation numberof CCTCC NO: M 2018860. The bacillus subtilis T8 has good cellulose degradation capability, can produce cellulase, and has strong endoglucanase activity, filter paper enzymatic activity, exonucleaseactivity and beta-glucosidase activity. The bacillus subtilis T8 can inhibit phomopsis, and an ethanol supernatant and a protein have inhibitory effects on the phomopsis. In addition, the bacillus subtilis T8 can inhibit the growth of escherichia coli, staphylococcus aureus, candida albicans, shigella dysenteriae and enterobacter faecalis, and has a development and utilization value in inhibitinghuman pathogenic bacteria. The invention also provides application of the bacillus subtilis T8 in producing cellulase and preventing kiwi fruit soft rot.

Description

technical field [0001] The invention belongs to the technical field of microorganisms, and in particular relates to a bacillus subtilis for inhibiting Phomopsis sp. and application thereof. Background technique [0002] Phomopsis (Phomopsis) is a large genus in the fungus of Deuteromycota, Coelospora, Coccoides, and Coccoidaceae. It contains more than 100 different species and can parasitize more than 70 different families. plant. The pathogenic bacteria of this genus are widely distributed in regions, causing serious diseases such as leaf blight, branch blight, stem rot, ulcer and fruit rot of plants, resulting in significant economic losses. For example, it is the soft rot of kiwi fruit after picking. The pathogenic bacteria of kiwifruit branch blight, peach stiff buds, and plantain panicle blight. [0003] At present, the prevention and control of Phomopsis mainly adopts chemical pesticides, such as: Yang Tingmi etc. adopt 8 kinds of pesticides to indoor toxicity and f...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12N9/42A01P3/00A61P31/04A61P31/10C12R1/125
CPCY02A50/30
Inventor 王金昌靳亮关丽梅郭燕
Owner INST OF MICROBIOLOGY JIANGXI ACADEMY OF SCI
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