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Mutant capable of raising cyclic lipopeptides antibiotic output by using RNA polymerase mutation, its preparation method and its application

An RNA polymerase and mutant technology, which is applied in the field of RNA polymerase mutant screening, can solve problems such as low yield, and achieve the effect of stabilizing physiological and biochemical characteristics and genetic traits

Inactive Publication Date: 2012-06-13
NANJING UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Such antibacterial substances are often produced in the stable growth period of microorganisms, and the low yield is the main reason restricting their development as biopesticides

Method used

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  • Mutant capable of raising cyclic lipopeptides antibiotic output by using RNA polymerase mutation, its preparation method and its application
  • Mutant capable of raising cyclic lipopeptides antibiotic output by using RNA polymerase mutation, its preparation method and its application
  • Mutant capable of raising cyclic lipopeptides antibiotic output by using RNA polymerase mutation, its preparation method and its application

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Experimental program
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Embodiment Construction

[0027] 1. Screening of RNA polymerase mutants

[0028] Activation of the strain

[0029] Inoculate wild-type Bacillus CC09 with 1% inoculum in 4 mL / tube of LB liquid medium, activate and culture at 37°C, 120 rpm for 48 hours, and then perform secondary activation in 8 LB liquid cultures with the same inoculum amount and culture conditions culture medium for 24 hours.

[0030] Preparation of rifampicin-containing plates

[0031] Add the rifampicin solution to the LB solid medium at 50-60°C to a final concentration of 50 μg / mL, which is the rifampicin-containing plate.

[0032] Will The secondary activation seed solution (8 test tubes) prepared in , was evenly spread on the plate containing rifampicin in the amount of 200 μL / dish, and the colonies were observed after 2 days of cultivation in the dark at 37°C, which was the possible mutation of RNA polymerase A total of 78 mutants were screened.

[0033] 2. Determination of the content of high-yield mutants and their cy...

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Abstract

The present invention belongs to the field of a biological technique and relates to a RNA polymerase mutant of Bacillus spp CC09 bacterial strain of high-yield antimycotics cyclic lipopeptides substances screened by using a LB plate containing rifampin (50 mug / ml). The mutation site of the mutant is on any position of 485-490 in a RNA polymerase beta sub-unit amino acid sequence, Compared with wild type CC09 bacterial strain, the output of antimycotics cyclic lipopeptides antibiotics Iturin and Surfactin produced by the mutant can be increased by 100%-300%, the antifungal activity is increased by 20-60%, and the mutant can be used for preventing and treating a plurality of crop diseases.

Description

technical field [0001] The invention belongs to the technical field of biological pesticides, and in particular relates to the screening and application of RNA polymerase mutants of bacillus CC09 strains producing antifungal cyclolipopeptide antibiotics. Background technique [0002] With the improvement of people's quality of life requirements and the various crises faced by the environment and ecology, the biggest challenge facing agricultural production is how to find environmentally friendly substitutes for chemical pesticides in the future to resist agricultural diseases. , biopesticides derived from beneficial microorganisms are the most promising substitutes for such products (Marc Ongena et al., Cell, 2007, 16(3): 115-125). Existing research on microbial pesticides mostly focuses on bacillus, and related products have appeared, such as the products SerenadeTM and Sonata AS of Agraquest Company in the United States, which are derived from bacterial strains Bacillus su...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12N15/01C12P21/04A01N63/02A01P3/00C12R1/07
Inventor 刘常宏李慧薛雅蓉
Owner NANJING UNIV
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