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Plant resistance-related protein ATWRKY46 as well as encoding gene and application thereof

A gene and protein technology, applied in the application field of plant resistance-related protein ATWRKY46 and its coding gene, can solve the problems of increasing salicylic acid, increasing the synthesis level of salicylic acid, etc., and achieve the effect of enhancing disease resistance

Active Publication Date: 2013-05-29
FUJIAN SANAN SINO SCI PHOTOBIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

So far, it has only been found that ETHYLENEINSENSITIVE3 (EIN3) and EIN3-LIKE1 can directly bind to the ICS1 promoter of the key SA biosynthesis gene, and negatively regulate the synthesis level of SA. It has not been found that ICS1 gene expression can be promoted by positively inducing the ICS1 promoter. proteins that increase the level of salicylic acid synthesis
In addition, studies have found that SAR Deficient 1 (SARD1) and CBP60g proteins are important factors in inducing ICS1 gene expression and SA synthesis, but failed to find more positive induction of ICS1 promoter to promote ICS1 gene expression, thereby increasing the level of salicylic acid synthesis protein

Method used

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  • Plant resistance-related protein ATWRKY46 as well as encoding gene and application thereof
  • Plant resistance-related protein ATWRKY46 as well as encoding gene and application thereof
  • Plant resistance-related protein ATWRKY46 as well as encoding gene and application thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0061] Embodiment 1, the acquisition of ATWRKY46 protein and its coding gene

[0062] 1. According to the existing NCBI database and literature, set a pair of primers as follows:

[0063] F1 (forward primer): 5'- TCTAGA CACATCTCCCACCAATCTCA-3' (underlined XbaI restriction recognition sequence);

[0064] R1 (reverse primer): 5'- CTCGAG TCGACCACAACCAATCCTGTC-3' (XhoI restriction recognition sequence is underlined).

[0065] 2. Genomic DNA was extracted from leaves of Arabidopsis thaliana ecotype Columbia.

[0066] 3. Using genomic DNA as a template, use the primer pair designed in step 1, and perform PCR amplification with PrimeSTAR HS high-fidelity enzyme from TaKaRa Company.

[0067] 4. The PCR amplification product is sequenced, as shown in sequence 2 of the sequence listing.

[0068] The protein shown in Sequence 1 of the Sequence Listing was named ATWRKY46 protein. The gene encoding the ATWRKY46 protein is named as the ATWRKY46 gene, and its genomic DNA is shown in ...

Embodiment 2

[0069] Embodiment 2, the cloning of each gene and the construction of its recombinant expression vector

[0070] 1. Acquisition of ATWRKY46 gene and construction of recombinant plasmid 326-ATWRKY46-FLAG

[0071] 1. Genomic DNA was extracted from leaves of Arabidopsis thaliana ecotype Columbia.

[0072]2. Using the genomic DNA in step 1 as a template, use the primer pair composed of F1 and R1 to carry out PCR amplification under the action of TaKaRa's PrimeSTAR HS high-fidelity enzyme to obtain PCR amplification products. The agarose gel electrophoresis of the PCR amplification product is shown in figure 1 (M represents a nucleotide marker, which is DL2000 from TaKaRa Company).

[0073] 3. The PCR amplification product of step 2 was double-digested with restriction endonucleases XbaI and XhoI, and the digested product was recovered.

[0074] 4. Digest the 326-FLAG expression vector with restriction endonucleases XbaI and XhoI, and recover the vector backbone of about 3827bp...

Embodiment 3

[0096] Embodiment 3, the application of ATWRKY46 gene in inducing ICS1 gene promoter (ProAtICS1) to start gene expression

[0097] 1. Transient expression of recombinant plasmids in Arabidopsis leaf protoplasts

[0098] The recombinant plasmid 326-ATWRKY46-FLAG and recombinant plasmid 326-Pro constructed in Example 2 AtICS1 ::GFP co-transformed Arabidopsis protoplasts (recombinant plasmid 326-T 7 -FLC is used as a negative control of the recombinant plasmid 326-ATWRKY46-FLAG; the 326-FLAG expression vector is used as another negative control of the recombinant plasmid 326-ATWRKY46-FLAG), the specific steps are as follows:

[0099] 1. Germinate Colombian ecotype Arabidopsis seeds on MS medium, transplant them into soil when the roots grow to 1-3 cm, and cultivate them in a greenhouse at 23° C. (12 hours of light per day, light intensity of 150 μE).

[0100] 2. Add 20ml of double distilled water to a 90mm petri dish, then add 1.82g of D-mannitol and dissolve it.

[0101] 3. T...

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Abstract

The invention discloses a plant resistance-related protein ATWRKY46 as well as an encoding gene and application thereof. The protein provided by the invention is named ATWRKY46 protein, which is from Arabidopsis thaliana and is any one of the following: (a) the protein formed by amino acid sequences as shown in a sequence 1; and (b) the protein which is derived from the sequence 1 in a way that the amino acid sequences as shown in the sequence 1 are subjected to replacement and / or deletion and / or addition of one or a plurality of amino acid residues, and has any function of the following: inducing a starting function of an ICS1 gene promoter, and inducing a starting function of a PR1 (pathogenesis-related protein) gene promoter, and the protein which is related to synthesis of salicylic acid in a plant and plant resistance and is derived from the sequence 1. The ATWRKY46 protein provided by the invention can increase disease resistance of the plant through regulating and controlling the synthesis of the salicylic acid in the plant. The invention is of great value to cultivate disease-resistant plants.

Description

technical field [0001] The invention relates to a plant resistance-related protein ATWRKY46 and its coding gene and application. Background technique [0002] Plants often suffer from various diseases in the process of cultivation and production. Some diseases pose a great threat to plant growth and seriously damage the yield and quality of crops. For plant diseases, at present, the main measures are to control them with drugs. Recently, people have begun to use abiotic inducers to induce plant disease resistance, and have been widely used in many important crops such as tobacco, potato, tomato, cucumber, bean, and rice. Although the effect of this measure is ideal, it is costly and pollutes the environment. Therefore, there is an urgent need to develop new biological means to improve the plant's own disease resistance. [0003] Systemic acquired resistance is a plant defense response that is induced when plants are attacked by pathogens and pests and can quickly spread t...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/415
Inventor 金京波蔡斌张强
Owner FUJIAN SANAN SINO SCI PHOTOBIOTECH CO LTD
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