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Cloning and application of arabidopsis oidium disease resistance suppressor gene EDTS3

A technology for suppressing genes and Arabidopsis thaliana, applied in the fields of application, genetic engineering, plant genetic improvement, etc., can solve the problem of low resistance and achieve the effect of improving disease resistance

Inactive Publication Date: 2012-07-18
INST OF GENETICS & DEVELOPMENTAL BIOLOGY CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Basic resistance is characterized by a broad spectrum of resistance, but usually to a low degree

Method used

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  • Cloning and application of arabidopsis oidium disease resistance suppressor gene EDTS3
  • Cloning and application of arabidopsis oidium disease resistance suppressor gene EDTS3
  • Cloning and application of arabidopsis oidium disease resistance suppressor gene EDTS3

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] Example 1 Analysis of the edts3 mutant suppressing disease resistance phenotype

[0035] 1. The edts3 / edr2 mutant inhibits the powdery mildew resistance and powdery mildew-induced cell death of the edr2 mutant

[0036] (1) Materials and methods

[0037] edts3 / edr2 (wherein edts3 is enhanced disease resistance two suppressor 3) is a mutant edr2 (4) that can suppress edr2 powdery mildew bacteria obtained by screening from the mutagenized population by using 0.03% EMS (purchased from sigma company) Mutants with enhanced resistance. Then a mutant was named edts3 / edr2.

[0038] First, the seeds of mutant edr2, edts3 / edr2 and wild-type col-0 (purchased from Arabidopsis Biological Resource Center (ABRC)) were sown on MS medium (commercially purchased from PhytoTechnology Laboratories), and vernalized at 4°C for 2-3 days , transferred to a plant growth chamber under 9h light / 15hdark light conditions. About 7-10 days, move the seedlings to the soil. After 4-6 weeks of growt...

Embodiment 2

[0041] Embodiment 2 Obtaining of EDTS3 gene

[0042] 1. Isolation of EDTS3 gene by map-based cloning

[0043] We isolated the EDTS3 gene using map-based cloning. The specific method is: the mutant edts3 / edr2 is crossed with the ecotype Lansberg (commercially purchased from ABRC), and the obtained F1 generation is self-crossed to generate the F2 generation. In the F2 generation, a single plant with a wild-type phenotype of the edr2 mutation was selected and then selfed to generate the F3 generation. Wild-type individuals were selected from each line of the F3 generation, and by PCR method and using chromosome positioning markers evenly distributed on the five chromosomes (http: / / signal.salk.edu / genome / SSLP_info / SSLPordered.html), the The gene was initially located at the front end of the broken arm of the first chromosome. Subsequently, about 3000 F3 generation individual plants were used to fine-map the gene in the 40kb region between F9H16 and F26F24 (8)( figure 2 ). Th...

Embodiment 3

[0046] Example 3 Functional Verification of EDTS3 Gene

[0047] In order to verify whether the resistance inhibition of edts3 mutation to edr2 mutant powdery mildew is caused by the mutation of At1g21327 base, we constructed a genetic transformation vector of At1g21327 with its own promoter, and transformed double mutant edts3 / edr2 plants by flower infection , in the T1 generation, the transgenic line recovered the phenotype of the edr2 mutant.

[0048] We used the wild-type col-0 DNA as a template, and used a pair of PCR primers F: 5′-AACTGCAGAAGTGGTGCTCACGTTGTTGTATTT-3′ and R: 5′-AACTGCAGAAGTGGTGCTCACGTTGTTGTATTT-3′ to amplify the 5.7kb At1g21327 gene with KpnI and PstI fragment.

[0049] PCR reaction system

[0050] DNA: 2.0ul

[0051] 10xbuffer: 5.0ul

[0052] dNTPs (2.0mM): 5.0ul

[0053] MgSO4 (25mM): 2.0ul

[0054] Primer F (10uM): 1.6ul

[0055] Primer R (10uM): 1.6ul

[0056] KOD Taq (1U): 1.0ul

[0057] H2O: 31.8ul

[0058] PCR reaction program: pre-denatur...

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Abstract

The invention relates to separated cloning, functional verification and application of arabidopsis oidium disease resistance suppressor gene EDTS3, and further relates to a mutant gene of the EDTS3 gene, protein encoded by the EDTS3 gene and application of the EDTS3 gene.

Description

technical field [0001] The application relates to the technical field of plant genetic engineering, in particular to the cloning, functional verification and application of a positive regulatory factor for powdery mildew resistance in the model plant Arabidopsis thaliana. Background technique [0002] Powdery mildew is a plant disease caused by fungi in the Powdery mildew family. When the plant is infested with disease, it will produce a large amount of visible white powder composed of mycelium, conidiophores and conidia, hence the name. Powdery mildew is widespread all over the world, especially in dicotyledonous plants. It also infects a variety of grasses, such as wheat, barley, oats and various pastures. For example, wheat powdery mildew is caused by the specialized parasitic fungus Erysibacter graminearum, and it is one of the main diseases affecting wheat production. In my country, wheat powdery mildew has been epidemic for several times, which caused great economic...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/29C07K14/415A01H5/00
Inventor 武广珩唐定中
Owner INST OF GENETICS & DEVELOPMENTAL BIOLOGY CHINESE ACAD OF SCI
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