WRKY45 gene of paddy rice, preparation method and application

A gene and rice technology, applied in the field of agricultural biology, can solve the problems that the WRKY45 gene function has not been truly revealed, and the gene function has not been elucidated, so as to achieve the effect of improving disease resistance and drought resistance and broad application prospects

Inactive Publication Date: 2007-07-25
XISHUANGBANNA TROPICAL BOTANICAL GARDEN CHINESE ACAD OF SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] But, before the present invention, the function of WRKY45 gene has not been revealed really yet; The research about WRKY45 gene improves

Method used

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  • WRKY45 gene of paddy rice, preparation method and application
  • WRKY45 gene of paddy rice, preparation method and application
  • WRKY45 gene of paddy rice, preparation method and application

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Effect test

Embodiment 1

[0034] Embodiment 1--The cloning of WRKY45 cDNA

[0035]Total RNA was extracted from rice seedlings using the modified method of Logemann et al. (1987). According to the procedure of the cDNA synthesis and library construction kit (BD Clontech), a cDNA library induced at low temperature (4° C.) for 2 h was constructed. According to the nucleotide sequence of the rice WRKY45 gene, corresponding primers are designed, and RT-PCR amplification is performed to obtain part of specific gene fragments. Subsequently, the RT-PCR amplification product was cloned into the pMD18-T vector (Dalian TaKaRa Company), and the amino acid sequence was sequenced and predicted. Finally, the obtained partial DNA fragment of the specific rice WRKY45 gene was used as a probe to screen the rice cDNA library to obtain the full-length cDNA of WRKY45, as shown in the sequence table.

Embodiment 2

[0036] Example 2--Screening of Highly Expressed WRKY45 Transgenic Plants

[0037] The obtained full-length cDNA of WRKY45 was cloned into pOCA30. The constructed plasmid was transformed into Agrobacterium LBA4404, and the living plant Agrobacterium flower dipping method was used for transformation, and the obtained transgenic seeds were obtained by using a method containing kanamycin (50 μg ml -1 ) MS medium for screening to obtain T 1 Transgenic plants were further confirmed by northern blot analysis. The test results are shown in Figure 1.

Embodiment 3

[0038] Embodiment 3--disease resistance analysis

[0039] Highly expressed WRKY45 and wild-type Arabidopsis plants grown for 4 weeks, Arabidopsis leaves injected with Pseudomanassyringae pv.Tomato strain DC3000, the bacterial solution concentration is OD 600 = 0.001. After culturing for 72 hours, samples were taken and plated, and the number of colonies was counted for analysis. The test results are shown in Figure 2.

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Abstract

This invention discloses a method for preparing rice gene WRKY45 and its applications. This invention successfully separates gene WRKY45 from rice, determines its nucleotide sequence, and deduces its amino acid sequence. Gene functional analysis shows that rice gene WRKY45 can effectively improve the disease and drought resistance of Arabidopsis thaliana, rice, economic crops, horticultural plants, and pasture grass, thus has wide potential applications.

Description

technical field [0001] The invention relates to agricultural biotechnology mainly based on transgenic plant technology, and more specifically, the invention relates to improving plant broad-spectrum disease resistance and drought resistance by using rice WRKY45 gene. Background technique [0002] The WRKY gene family of transcriptional regulators is a plant-specific super gene family, which contains highly conserved amino acid sequences WRKYGQK and CX 4-5 CXnHXH / C (X is any amino acid) at the N-terminal of its protein, and belongs to zinc finger transcriptional regulators . WRKY protein can specifically bind the DNA-specific sequence TTGACC (W box) of the target gene promoter, thereby regulating gene expression (EulgemT. et al., 2000, Trends Plant Sci., 5:199-206; Yu et al. 2001, The Plant Cell, 13:1527-1539). [0003] It is currently known that the main biological function of the WRKY gene family is to regulate signal transduction pathways in plant organic integration, bi...

Claims

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Application Information

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IPC IPC(8): C12N15/29C07K14/415C12N15/82
Inventor 余迪求仇玉萍
Owner XISHUANGBANNA TROPICAL BOTANICAL GARDEN CHINESE ACAD OF SCI
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