Pathogenicity gene MoSNT2 from magnaporthe oryzae and application thereof

A technology of rice blast fungus and pathogenicity, applied in the field of microbial genetic engineering, can solve the problems of rice blast damage and lack of effective control methods, and achieve the effect of controlling pathogenicity and epidemic

Pending Publication Date: 2017-09-26
SICHUAN AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] In view of the serious damage of rice blast currently existing and the lack of effective control methods, the purpose of the present invention is to provide a pathogenic protein derived from rice blast fungus

Method used

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  • Pathogenicity gene MoSNT2 from magnaporthe oryzae and application thereof
  • Pathogenicity gene MoSNT2 from magnaporthe oryzae and application thereof
  • Pathogenicity gene MoSNT2 from magnaporthe oryzae and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] Example 1: Isolation and cloning of the MoSNT2 gene

[0043] (1) The SNT2 protein plays an important role in the oxidative stress response of yeast (Baker et al., Molecular and Cellular Biology, 2013, (33) 19:3735–3748), but the function of this protein in Magnaporthe grisea is unknown. There is a certain similarity between the protein sequences of yeast and Magnaporthe oryzae, so first search the yeast SNT2 protein sequence from the yeast genome database (http: / / www.yeastgenome.org / ), and then use blastP to search the genome of Magnaporthe oryzae database( http: / / www.broadinstitute.org / annotation / genome / magnaporthe_ grisea / MultiHome.html ) was searched for homologous sequences, and a predicted protein MGG_04421 of Magnaporthe grisea was obtained, which was named MoSnt2. The amino acid sequence of the protein is shown in SEQ ID NO: 1.

[0044] (2) Cloning of the MoSNT2 gene: for cloning the cDNA sequence of MoSNT2, first use the RNeasy Plant MiniKit kit (Qiagen Comp...

Embodiment 2

[0048] Example 2: Quantitative RT-PCR detection of MoSNT2 gene expression levels at different developmental stages

[0049] (1) Preparation of media and solutions:

[0050] (1) CM liquid medium: its composition and proportion are: D-Glucose 10g, Peptone 2g, YeastExtract 1g, Casamino Acids 1g, 20×Nitrate Salts * 50ml, Vitamin Solution # 1ml, TraceElements & 1ml, add ddH 2 0 to 1000ml, and adjust the pH value to 6.5 with 1mol / L NaOH solution.

[0051] (2) CM agar plate: its composition and proportion are: D-Glucose 10g, Peptone 2g, YeastExtract 1g, Casamino Acids 1g, 20×Nitrate Salts * 50ml, Vitamin Solution # 1ml, TraceElements & 1ml, 15g agar powder, add ddH 2 O to 1000ml, and adjust the pH value to 6.5 with 1mol / L NaOH solution; sterilize with damp heat at 121°C for 20min.

[0052] (3) * 20×Nitrate Salts (1000ml) composition and proportion are: NaNO 3 120g, KCl 10.4g, MgSO 4 ·7H 2 O 10.4g, KH 2 PO 4 30.4g, add ddH 2 O to 1000ml, sterilize with damp heat at 12...

Embodiment 3

[0060] Example 3: Construction of Magnaporthe grisea MoSNT2 gene knockout mutant

[0061] Proceed as follows:

[0062] Adopt the Split-Marker knockout method (references: Kershaw et al., P Natl Acad Sci USA, 2009, 106 (37): 15967-15972) to construct the knockout mutant of gene MoSNT2 (see figure 2 ). The coding sequence of about 2.5kbp of the gene MoSNT2 was selected as the targeting site. During the gene knockout process, the homologous recombination event replaced the targeting site sequence with the hygromycin selection marker gene HYG, thereby forming a gene knockout mutant. The specific construction steps are as follows:

[0063] (1) Using the genomic DNA of the blast fungus wild-type Guy11 as a template, using SNT2-LF-For and SNT2-LF-Rev, or SNT2-RF-For / SNT2-RF-Rev as primers, DNA polymerase KOD- Plus (Kangwei Century Biotechnology Co., Ltd.) performed PCR amplification to obtain the left-wing LF (SEQ ID NO: 7) or right-wing RF fragment (SEQ ID NO: 8) of the MoSNT2 g...

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PUM

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Abstract

The invention discloses a pathogenic protein MoSNT2 derived from Magnaporthe grisea, the protein is composed of the amino acid sequence shown in SEQ ID No: 1; the invention also discloses the gene MoSNT2 encoding the pathogenic protein, the gene It consists of the nucleotide sequence shown in SEQ ID No:2. The invention also discloses the use of the above-mentioned gene or protein as a target for designing or screening antifungal drugs. The invention provides an important target gene or target protein for the control of rice blast. The drug developed by using the gene or protein as a target can destroy the gene expression or protein function of the rice blast fungus, and can effectively control the pathogenicity and prevalence of the rice blast fungus.

Description

technical field [0001] The invention belongs to the field of microbial genetic engineering, and specifically relates to a pathogenic gene MoSNT2 derived from rice blast fungus, and also relates to the use of the gene. Background technique [0002] Rice blast (having another name called rice fever) is one of the important diseases of rice, which can cause a substantial reduction in production, and when serious, reduce production by 40% to 50%, or even fail to harvest. Rice blast occurs in all rice regions in the world, and most of them occur on leaves and nodes, especially the early and severe occurrence of panicle blast or node blast, which can cause white ears and even extinction. The pathogen that causes rice blast is Magnaporthe oryzae, a semi-living parasitic fungus (Magnaporthe oryzae). It has the characteristics of many physiological races, rapid reproduction and variation, and strong prevalence, which poses a huge threat to rice production. In order to ensure the saf...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/37C12N15/31C12R1/645
CPCC07K14/37
Inventor 陈学伟贺闽许有嫔陈金华李伟滔王静
Owner SICHUAN AGRI UNIV
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