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Bacillus subtilis lipo-peptide biological pesticide and use

A technology of Bacillus subtilis and bio-pesticides, applied in the biological field, can solve problems such as application limitations and destruction of red blood cells

Inactive Publication Date: 2004-12-15
NANJING AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] So far, the research targets of lipopeptide compounds are mainly focused on medical applications, but the application of lipopeptide compounds in medicine is limited due to the effect of destroying red blood cells. Molecular modification of lipopeptide compound enzyme genes It is still a hotspot in biochemistry and molecular biology research in the world to create new active compounds that can be used in medicine
So far, the research work on lipopeptides has been limited to the research of molecular biology in the laboratory. There are no literature and patent reports to use industrial fermentation methods to produce lipopeptides from Bacillus subtilis, and use the lipopeptides to treat plant diseases and insect pests. report on prevention

Method used

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  • Bacillus subtilis lipo-peptide biological pesticide and use
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  • Bacillus subtilis lipo-peptide biological pesticide and use

Examples

Experimental program
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Effect test

Embodiment 1

[0024] 1. The production method of Bacillus subtilis B3 strain lipopeptide biological pesticide, including

[0025] Prepare 20L of fermentation medium in a 30L fermentor (NLF, Bioengineering AG) in Switzerland Bioengineering AG: 20g sucrose, 1g yeast extract, 0.5g sodium glutamate, 0.5gMgSO per litre 4 , 0.5gKCl, 1gKH 2 PO 4 , 0.15mgFe 2 SO 4 ·6H 2 O, 5.0mgMnSO 4 ·H 2 O, 0.16mgCuSO 4 ·5H 2 O, adjust the pH range to 7.0, add water to 1L;

[0026]The seed bacteria were cultured with shaking for 12 hours in a shake flask, and 400 mL of seed bacteria was added to the fermentor at 2% of the volume of the fermentation medium. The temperature was 35°C, the rotation speed was 500r / min, the dissolved oxygen was 30%, and the fermentation was 30h. After the fermentation, it was collected by centrifugation. Add 6mol / L HCl to the supernatant to adjust the pH to 2.0, and collect the precipitate by centrifugation. The wet weight is about 142g. After adding methanol at 1 times its mass, adjust th...

Embodiment 2

[0033] 1. The production method of Bacillus subtilis B213 strain lipopeptide biological pesticide, including

[0034] Prepare 20L of fermentation medium in a 30L fermenter (NLF, Bioengineering AG) in Switzerland Bioengineering: 15g glucose, 0.5g yeast extract, 0.5g sodium glutamate, 0.5gMgSO per liter of culture broth 4 , 0.5gKCl, 1gKH 2 PO 4 , 0.15mgFe 2 SO 4 ·6H 2 O, 5.0mgMnSO 4 ·H 2 O, 0.16mgCuSO 4 ·5H 2 O, adjust the pH range to 7.0, add water to 1L;

[0035] The seed bacteria were cultured in a shake flask for 12 hours. Add 400 mL of seed bacteria to the fermentation tank at 2% of the fermentation medium volume. The temperature was 35~37℃, the rotation speed was 500r / min, the dissolved oxygen was 30%, and the fermentation was 30h. After the fermentation, it was collected by centrifugation. Supernatant, add 6mol / L HCl to the supernatant to adjust the pH to 2.0, centrifuge to collect the precipitate, the wet weight is about 160g, add methanol at 1 times its mass, adjust the pH ...

Embodiment 3

[0040] Different Bacillus subtilis strains, using different medium components, using the method and conditions of Example 1, can also obtain lipopeptide biological pesticides through fermentation production and quality inspection, and the yield of lipopeptide biological pesticides can be 10~ 35g (see Table 2), the field application method and effect are the same as in Example 1, which can be achieved by those skilled in the art.

[0041] See J. Vater, XWGao, G. Hitzeroth, C. Wilde and P. Franke. "Whole cell" for the strains B3, B213, 43, 49, ATCC6633, OKB105 and C1 used in the above examples and the pesticide MALDI-TOF-MS detection method. "-Matrix-Assisted Laser DesorptionIonization-Time of Flight-Mass Spectrometry, an Emerging Technique for Efficient Screening of Biocombinatorial Libraries of NaturalCompounds-Present State of Tesearch. Combinatorial Chenistry & High Throughput Screening, 2003, 6, 557-567.

[0042] Table 1 The effects of lipopeptide bio-pesticides on crop disease...

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Abstract

The present invention relates to biological bacillus lipopeptide pesticide and its application, and belongs to the field of biotechnology. The lipopeptide pesticide is prepared through fermentation culture in culture medium of glucose, cane sugar or corn slurry at temperature of 30-37 deg.c, rotation speed of 300-1000 rpm and dissolved oxygen of 30% for 30-36 hr; centrifugation to collect supernatant, adding HCl to regulate pH to 2.0-2.5, centrifugal extraction to collect supernatant, methanol or ethanol extraction, merging extracted liquid, and vacuum drying. The present invention is used in the green prevention and control of pests and diseases of field.

Description

(1) Technical field [0001] The Bacillus subtilis lipopeptide biological pesticide and application of the invention belong to the field of biotechnology. It is specially used to prepare lipopeptide compounds from Bacillus subtilis for the pollution-free prevention and control of plant diseases and insect pests and increase crop yield. (2) Background technology [0002] The lipopeptide compound is a natural antibacterial substance produced by Bacillus subtilis with potential plant disease control and medical application value. It includes surfactin (surfactin), iturins (iturins) and fengycin. Among them, surfactin is not only a highly active biosurfactant, but also a bioactive substance with a wide range of uses. It exhibits antiviral, antitumor and antimycoplasma activities, as well as good performance in petroleum recovery and demulsification. And iturins can affect the surface tension of the cell membrane, resulting in the formation of micropores, K + The leakage of other import...

Claims

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Application Information

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IPC IPC(8): A01N63/02
Inventor 高学文刘俊王金生
Owner NANJING AGRICULTURAL UNIVERSITY
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