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Effector R1YBX6 related to rhizoctonia solani pathopoiesia and its encoding gene

A technology of R1YBX6 and Rhizoctonia solani, applied in the direction of genetic engineering, plant gene improvement, application, etc., can solve the problem of not getting normal plants

Inactive Publication Date: 2018-04-24
YUNNAN AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] Gene transfer using the Ti plasmid in Agrobacterium as a vector has the characteristics of easy operation, but all the time, this transfer system is only suitable for dicotyledonous plants, and there are few successful examples of monocotyledonous plants. Ti plasmids can transform mature rice embryos, and T-DNA has been integrated into the cell chromosome and expressed, but no normal plants have been obtained from the transformed tissues; the existing ones are obtained in indica rice, japonica rice and bare rice by optimizing various factors. In addition, using the young embryos and young panicles of indica rice varieties Aoqingzhan, Zaosizhan and Qishanzhan as explants, explore the transfer of GUS gene mediated by Agrobacterium in indica rice

Method used

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  • Effector R1YBX6 related to rhizoctonia solani pathopoiesia and its encoding gene
  • Effector R1YBX6 related to rhizoctonia solani pathopoiesia and its encoding gene
  • Effector R1YBX6 related to rhizoctonia solani pathopoiesia and its encoding gene

Examples

Experimental program
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Effect test

Embodiment 1

[0023] Example 1. Introduction of secreted protein gene into binary vector pBI121:

[0024] The vector used is pBI121, and the insertion site is XbaI / BamHI;

[0025] (1) Target fragment amplification: design primers according to the sequence, and amplify a sufficient amount of the target product by PCR; the first round of PCR reaction uses pfu polymerase; reaction system 50ul; DNA 1ul; primer MIX 10ul; 1ul each primer; dNTP 1ul; 10X pfu Buffer 5ul; Pfu 0.4ul, dilute to 50ul;

[0026] PCR reaction procedure: the first step, the reaction temperature is 95°C, the reaction time is 3min; the second step, the reaction temperature is 94°C, the reaction time is 30s; the third step, the reaction temperature is 55°C, the reaction time is 30s; Four steps, the reaction temperature is 72°C, the reaction time is 60s; the fifth step, the reaction temperature is 72°C, the reaction time is 6min; the second step to the fourth step cycle 22 times;

[0027] 1) Using the first-round PCR product ...

Embodiment 2

[0037] Embodiment 2, the culture transformation of Agrobacterium tumefaciens:

[0038] The Cacl2 freeze-thaw method was used to transfer the plasmids into EHA105 Agrobacterium competent cells;

[0039] Add 0.5-1 μg of plasmid DNA to a centrifuge tube containing 200 μL of EHA105 Agrobacterium competent cells, mix gently; place in ice bath for 30 minutes, then transfer to liquid nitrogen and freeze for 5 minutes; quickly place in a 37°C water bath 5 minutes;

[0040] Add 600 μL of LB liquid medium, control the rotation speed to 200r / min, and shake at 28°C for 4h;

[0041] Spread the activated Agrobacterium on the LB solid medium of 50mg / L kanamycin and 20mg / L rifampicin, and culture it upside down at 28°C for 2-3 days;

[0042] Select a single clone in 500 μL LB liquid medium containing 50 mg / L kanamycin and 20 mg / L rifampicin, control the temperature at 28 ° C, rotate at 200 r / min, and shake for 10 h;

[0043] Pipette 10ul of Agrobacterium culture solution into 500ul LB liqu...

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Abstract

The invention discloses an effector R1YBX6 related to rhizoctonia solani pathopoiesia and its encoding gene. The invention relates to the effector R1YBX6 related to rhizoctonia solani pathopoiesia andfunction and sequence of its encoding gene. The transgenic instantaneous expression shows that the R1YBX6 refers to one of multigene of rhizoctonia solani pathopoiesia; the fining of the gene contributes to the molecular mechanism study for learning rhizoctonia pathopoiesia.

Description

technical field [0001] The invention relates to an effector R1YBX6 related to the pathogenicity of Rhizoctonia solani and its coding gene, belonging to the field of biotechnology. Background technique [0002] Rice sheath blight is caused by Rhizoctonia soalni, which is a soil-inhabiting fungus widely distributed in rice-producing areas around the world; it not only infects a wide range, but also often causes a variety of plant diseases , and has strong saprophytic competition ability in soil and long survival time; therefore, it is considered to be one of the most destructive soil-borne plant pathogens; An important disease in the region; up to now, some studies have been done on the molecular mechanism of disease resistance related to this disease at home and abroad, but due to the lack of relevant data on the pathogenic molecular mechanism of Rhizoctonia solani, no breakthrough has been made in disease-resistant breeding. Progress; there are few studies on the effector p...

Claims

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Application Information

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IPC IPC(8): C07K14/37C12N15/31C12N15/82C12N5/10
CPCC07K14/37C12N15/8205
Inventor 李成云董文汉包文静
Owner YUNNAN AGRICULTURAL UNIVERSITY
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