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Verticillium dahliae adp-atp carrier protein disease course key target gene and its interference carrier and application

A technology of Verticillium dahliae and carrier protein, which is applied in the key target gene of ADP-ATP carrier protein disease course of Verticillium dahliae and its interference carrier and application field, which can solve the problems of less research and improve disease resistance Effect

Active Publication Date: 2016-08-31
THE INST OF BIOTECHNOLOGY OF THE CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] At present, the research on AACP mainly focuses on its relationship with energy metabolism, and there are few studies in fungi, especially its relationship with pathogenicity

Method used

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  • Verticillium dahliae adp-atp carrier protein disease course key target gene and its interference carrier and application
  • Verticillium dahliae adp-atp carrier protein disease course key target gene and its interference carrier and application
  • Verticillium dahliae adp-atp carrier protein disease course key target gene and its interference carrier and application

Examples

Experimental program
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Effect test

Embodiment 1

[0048] Example 1 Screening of ADP-ATP carrier protein disease course key target genes of Verticillium dahliae

[0049] 1. Experimental method

[0050] 1.1 Construction of VIGS interference vector

[0051] In order to obtain the fragment with the best interference effect on the target gene, according to the cDNA sequence of Verticillium dahliae ADP, ATP carrier protein (VDAG_07535.1), design specific primers (Table 1), with BamH I and EcoR I at both ends of the primers Restriction site, the target gene was subjected to PCR amplification, and the PCR product was detected by 1% agarose gel electrophoresis. Then, the recovered fragments were ligated into TRV2 vectors, verified by enzyme digestion and sequencing, and the verified plasmids were transformed into Agrobacterium GV3101.

[0052] Table 1 Primer information of different segments of AACP

[0053]

[0054] Note: The bolded part is the restriction site.

[0055] 1.2 VIGS transformation method

[0056] Agrobacterium s...

Embodiment 2

[0072] Construction and plant transformation of embodiment 2Gateway interference vector

[0073] 1. Experimental method

[0074] 1.1 Construction of stable genetic vector

[0075] In order to obtain stably inherited Nicotiana benthamiana containing the target gene dsRNA, according to the change of the tobacco disease index of transient transformation in Example 1, three DNA segments were selected (the nucleotide sequences are respectively SEQ ID No.1, SEQ ID No. 3. Shown in SEQ ID No.5), redesign the primers containing some BP sites at both ends, and then use attb primers to amplify (Table 3), which is used for the construction of Gateway interference vectors. According to the BP reaction, the target sequence was ligated into pDONR207; then it was constructed into pK7GWIWG2(I),0 by the LR reaction. Finally, the constructed vector was transformed into Agrobacterium LBA4404.

[0076] Table 3 Gateway interference primer information

[0077]

[0078] 1.2 Plant Transformatio...

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Abstract

The invention discloses a verticillium dahliae ADP-ATP carrier protein (AACP) pathogenesis key target gene as well as an interference vector and application thereof, belonging to the fields of cloning and application of a verticillium dahliae pathogenesis key gene. According to the invention, 3 sections of pathogenesis key target genes which are capable of significantly reducing disease index in a plant are screened from an AACP gene by virtue of a host induced gene silencing technology, and the 3 sections of target segments are further used for constructing a Gateway interference vector, so as to obtain transgenic tobacco which is significantly high in pathogenic bacterium resistance and stable in genetic performance. Through disease index and fungal biomass analysis as well as transcriptional level detection of the target gene, an optimum interference section of the AACP gene is finally screened and obtained. The verticillium dahliae ADP-ATP carrier protein pathogenesis key target gene and the RNA interference vector disclosed by the invention are applicable to improving disease resistance of plants to verticillium dahliae and breeding new varieties of transgenic plants resisting verticillium dahliae.

Description

technical field [0001] The present invention relates to the key target gene of the disease course of the ADP-ATP carrier protein of Verticillium dahliae, and also relates to an RNA interference carrier containing the key target gene of the disease course of the ADP-ATP carrier protein. The present invention also relates to the ADP- The application of the ATP carrier protein disease process key target gene or interference carrier in improving plant resistance to Verticillium dahliae belongs to the field of cloning and application of key gene disease process of Verticillium dahliae. Background technique [0002] Cotton verticillium wilt was first discovered in the United States in 1914 and has become a worldwide disease. The pathogen is Verticillium dahliae. Verticillium dahliae belongs to Deuteromycotina, Moniliaceae, and Verticillium. It not only seriously endangers the fiber quality and lint yield of cotton, but also harms 40 families. 660 kinds of plants, including 184 ki...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/31C12N15/63C12N5/10A01H5/00
Inventor 苏晓峰齐希梁郭惠明程红梅
Owner THE INST OF BIOTECHNOLOGY OF THE CHINESE ACAD OF AGRI SCI
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