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Application of elicitor protein derived from sarocladium fungi in prevention and control of cassava bacterial wilt
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A kind of elicitor, the technology of Scleroderma
Active Publication Date: 2020-09-29
ENVIRONMENT & PLANT PROTECTION INST CHINESE ACADEMY OF TROPICAL AGRI SCI
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So far, many new protein elicitors have been discovered, but there is no report on the elicitor protein and its encoding gene that can induce plant disease resistance in the fungus of the genus Cladostratum
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Embodiment 1
[0042] Embodiment 1, the cloning of the coding gene of elicitor protein SbES and its cDNA
[0043]A high-efficiency biocontrol strain isolated from Brachiaria leaves in our laboratory was identified as Sarocladium brachiariae and named as Sarocladium brachiariae HND5 (Liu X, Guo Z, Huang G: Sarocladium brachiariae sp. nov., an endophytic fungus isolated from Brachiaria brizantha. Mycosphere 2017, 8:827-834.). The inventors of the present invention, on the basis of the whole genome sequencing data of the strain, compared and analyzed the elicitor sequences reported by relative fungi, found and identified the potential for improving plant resistance and inducing plant defense response activity. Elicitor protein gene.
[0044] The method for obtaining the full-length coding DNA of the elicitor protein gene of the present invention and the full-length cDNA fragment is as follows:
[0045] Using the total DNA extracted from the Sarocladium brachiariae HND5 strain as a template, d...
Embodiment 2
[0052] Embodiment 2, eukaryotic expression and purification of SbES protein
[0053] 1. Optimization of cDNA sequence based on codon preference of Pichia pastoris
[0054] In order to use Pichia pastoris X-33 strain for heterologous expression of SbES protein, according to the codon preference of Pichia pastoris, the signalpeptide and propeptide encoding genes of SbES protein were excluded, and the 316-1164 nucleotides of sequence 2 were Codon optimization was carried out with acid to obtain sequence 4, which is encoded from the protein shown in amino acid 106-387 of the amino terminal of sequence 1 in the sequence listing. Sequence 4 was synthesized by Beijing Huada Biotechnology Co., Ltd.
[0056] The primers SbES-PF and SbES-PR were paired, and the sequence 4 was used as a template, and the amplified fragment was connected to the T vector, named pMD18-T / SbESp; pMD18-T / SbESp was double-digested with EcoRI and KpnⅠ, ...
Embodiment 3
[0066] Example 3, Evaluation of the control effect of SbES protein on cassava bacterial wilt
[0067] Pathogen strain: Xanthomonas axonopodis strain XamGX11 (Construction and quality evaluation of the transposon library of cassava bacterial wilt [J]. Journal of Tropical Crops, 2013, 34(6): 1144-1148.) , the public can obtain the strain from the Institute of Environment and Plant Protection, Chinese Academy of Tropical Agricultural Sciences.
[0068] Cassava variety: SC10
[0069]Medicament: recombinant protein SbES (0.1 mg / mL), Artelin (3% oligosaccharide protein·3% Alternaria activator protein WP) (Institute of Plant Protection, Chinese Academy of Agricultural Sciences), and PBS Buffer as the control.
[0070] Chemical treatment: Adjust the concentration of the recombinant protein His-SbES to 0.1 mg / mL, spray 10 potted cassava seedlings for 3 times with an interval of 3 days, and treat with Alterin: Alterin (3% oligosaccharide protein·3% chain spacer Sporogenin WP) was ...
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Abstract
The invention discloses an application of an elicitorprotein derived from sarocladium fungi in prevention and control of cassava bacterial wilt. According to the application, the elicitorprotein derived from sarocladium fungi is used for inducing cassava to generate disease resistance to bacterial wilt or improving the disease resistance of cassava. The elicitorprotein is a protein with one ofthe following amino acid residue sequences: 1) an amino acid residue sequence shown as SEQ ID No.1 in a sequence table; and 2) a protein which is derived from SEQ ID No.1, obtained by substituting and / or deleting and / or adding one or more amino acid residues in the amino acid residue sequence of SEQ ID No.1 in the sequence table and having an elicitor protein function. Experiments prove that the protein elicitor SbES can induce cassava to generate or improve the resistance of plants to target pathogenic bacteria, reduce the morbidity and the disease index, provide a new way for improving the disease resistance of the plants, and have a wide application prospect in agricultural production.
Description
technical field [0001] The invention belongs to the field of molecular biology, and relates to the application of an elicitor protein derived from a fungus of the genus Brachia and its coding gene in preventing and treating cassava bacterial wilt. Background technique [0002] Protein elicitors are derived from some proteins secreted by bacteria and fungi, mainly including Harpin, Cryptogein, and Activator. By inducing the expression of plant defense-related genes and activating defense signaling pathways in plants, such as salicylic acid, etc., plants can be stimulated to acquire resistance, and plant self-immunity can be enhanced to resist the infection of pathogenic microorganisms. Many new protein elicitors have been discovered so far, but there is no report on the elicitor protein and its encoding gene that can induce plant disease resistance in the fungus of the genus Cladopa. SUMMARY OF THE INVENTION [0003] The high-efficiency biocontrol strain isolated from the ...
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