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Construction method and application of recombinant Streptomyces lydicus for inhibiting plant pathogenic fungi

The technology of a plant pathogenic fungus, Streptomyces lydida, is applied in the field of bioengineering, which can solve the problems of poor antibacterial ability, single antibacterial mechanism, and narrow antibacterial spectrum.

Active Publication Date: 2015-11-11
BEIJING ACADEMY OF AGRICULTURE & FORESTRY SCIENCES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, as a natural strain, the metabolites of a single strain are limited, and there will be a single antibacterial mechanism, poor antibacterial ability, and narrow antibacterial spectrum.

Method used

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  • Construction method and application of recombinant Streptomyces lydicus for inhibiting plant pathogenic fungi
  • Construction method and application of recombinant Streptomyces lydicus for inhibiting plant pathogenic fungi
  • Construction method and application of recombinant Streptomyces lydicus for inhibiting plant pathogenic fungi

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0106] Embodiment 1, construct recombinant Streptomyces lydidii

[0107] This example provides a protein and its coding gene derived from Paenibacillus polymyxa JZB120001CGMCC No.5563, and the protein is named RFRP. The amino acid sequence of RFRP is shown in SEQ ID No. 1 in the sequence listing, and the sequence of its coding gene is shown in nucleotides 14-1618 of SEQ ID No. 2 in the sequence listing.

[0108] 1. Construction of recombinant vector

[0109] Replace the NdeI and the EcoRI recognition site in pIB139 with the DNA fragment between the NdeI of SEQIDNo.2 in the sequence listing and the EcoRI recognition site (i.e. the DNA fragment shown in the 14th-1618th nucleotide of SEQIDNo.2 in the sequence listing) For the DNA fragment between the dots, other sequences of pIB139 remain unchanged, and a recombinant vector containing the RFRP gene is obtained, and the recombinant vector is named pIB139-RFRP. In pIB139-RFRP, the expression of RFRP gene is initiated by the eryth...

Embodiment 2

[0119] Embodiment 2, the ability of degrading lichen polysaccharide of Streptomyceslydicus (Streptomyceslydicus) AB01CGMCCNo.10707

[0120] The experiment was repeated three times, and the results of each repeated experiment were as follows:

[0121] Streptomyces lydicus (Streptomyceslydicus) AB01CGMCCNo.10707, Streptomyces lydicus (Streptomyces lydicus) A01 and Streptomyces lydicus (Streptomyces lydicus) A01 of embodiment 1 were respectively inoculated on the lichenin polysaccharide screening plate, cultivated for 2 days, and carried out Congo red staining experiment. The results showed that the lichen polysaccharide screening plate growing Streptomyceslydicus (Streptomyceslydicus) AB01CGMCCNo.10707 and Lidi Streptomyces lydicus (Streptomyceslydicus) AG01 all produced transparent circles, while the lichen polysaccharide screening plate growing Streptomyceslydicus (Streptomyceslydicus) A01 had no transparency The circle appears, and the transparent circle produced by the lich...

Embodiment 3

[0123] Embodiment 3, the antibacterial activity of Streptomyces lydicus (Streptomyceslydicus) AB01CGMCCNo.10707

[0124]1. Preparation of plant fungal disease inhibitors

[0125] Streptomyces lydicus (Streptomyceslydicus) AB01CGMCCNo.10707 of embodiment 1 was inoculated on Gao's No. 1 slant medium, cultivated for 7-10 days at 28°C, until it produced a sufficient amount of spores, scraped it with a sterile platinum ring 2-3 loops of spores were inoculated into 50ml seed medium in a 250ml Erlenmeyer flask, placed on a temperature-controllable shaker, and cultured for 24h-30h at a constant temperature of 200rpm / min (rotation radius: 13mm) at 28°C; Under the conditions, it is tapped in the fermentation medium in 10 500ml Erlenmeyer flasks (the amount of liquid in each bottle is 100ml), and the OD of each bottle is inoculated. 600 The values ​​are all 0.1; the shake flasks after inoculation were shaken and cultured at a speed of 240rpm / min (radius of rotation: 13mm) for 96 hours a...

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Abstract

The invention discloses a construction method and application of recombinant Streptomyces lydicus for inhibiting plant pathogenic fungi. The method for constructing recombinant Streptomyces lydicus comprises the following step: transforming coding gene of a protein into Streptomyces lydicus as a host bacterium to obtain the recombinant Streptomyces lydicus. The protein is a) or b): a) protein of which the amino acid sequence is SEQ ID NO.1 in the sequence table; and b) protein with same functions subjected to substitution and / or deletion and / or addition of one or more amino acid residues on the amino acid sequence disclosed as SEQ ID NO.1 in the sequence table. The experiment proves that the Streptomyces lydicus AB01CGMCC No.10707 and RFRP can be used for inhibiting plant pathogenic fungi and preparing inhibitors for plant pathogenic fungi.

Description

technical field [0001] The invention relates to a construction method and application of a recombinant Streptomyces lydides inhibiting plant pathogenic fungi in the field of bioengineering. Background technique [0002] my country is a large agricultural country based on planting. Many major diseases, such as gray mold, anthracnose, tomato leaf mold, watermelon wilt, wheat head blight, etc., have caused serious losses to agricultural production (Chen Jun et al., 2010 ). The economic loss can be reduced by about 30 billion yuan through the use of pesticides every year, but chemical control plays a major role. The amount of chemical pesticides used accounts for more than 80% of the amount of pesticides used, causing serious environmental pollution, pesticide residues, increased resistance, etc., seriously threatening human health and the environment. Therefore, biological control is an inevitable trend in the development of pesticides. Due to the high specificity of microorga...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/76C12N1/15A01N63/00A01N63/02A01P3/00C12R1/465
Inventor 吴慧玲刘伟成李锦锦刘霆董丹张涛涛卢彩鸽刘德文张殿朋
Owner BEIJING ACADEMY OF AGRICULTURE & FORESTRY SCIENCES
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