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Plant resistance related protein ANAC001, its encoding gene and application thereof

A gene and protein technology, applied to the plant resistance-related protein ANAC001 and its coding gene and application field, can solve the problems of increasing salicylic acid, increasing the level of salicylic acid synthesis, and so on, so as to achieve the effect of enhancing disease resistance

Active Publication Date: 2013-07-17
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 ANAC001, its encoding gene and application thereof
  • Plant resistance related protein ANAC001, its encoding gene and application thereof
  • Plant resistance related protein ANAC001, its encoding gene and application thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0061] Embodiment 1, the acquisition of ANAC001 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'-GC TCTAGA AAATTATTAGATATACCAAACCAG-3' (underlined XbaI restriction recognition sequence);

[0064] R1 (reverse primer): 5'-CG GGATCC GACCAACAAGAATGATCCAACT-3' (underlined BamHI restriction recognition sequence).

[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 ANAC001 protein. The gene encoding the ANAC001 protein is named ANAC001 gene, and its genomic DNA is shown in Sequ...

Embodiment 2

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

[0070] 1. Acquisition of ANAC001 gene and construction of recombinant plasmid 326-ANAC001-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 enzymes XbaI and BamHI, and the digested product was recovered.

[0074] 4. The 326-FLAG expression vector was double digested with restriction enzymes XbaI and BamHI, and the vector backbone of about 3827 bp was re...

Embodiment 3

[0096] Embodiment 3, the application of ANAC001 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-ANAC001-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-ANAC001-FLAG; the 326-FLAG expression vector is used as another negative control of the recombinant plasmid 326-ANAC001-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. Take ...

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Abstract

The invention discloses a plant resistance related protein ANAC001, its encoding gene and an application thereof. The protein provided by the invention is named as ANAC001 protein, comes from Arabidopsis thaliana, and is any one of the following proteins: (a) a protein formed by an amino acid sequence as shown in the sequence 1; (b) a protein which is derived from the sequence 1 by replacing and / or deleting and / or adding one or more amino acid residues in the amino acid sequence of the sequence 1, and has any one of the following functions: a function of inducing ICS1 gene promoter, a function of inducing PR1 gene promoter, a function of being related with salicylic acid synthesis in plant and a function of being related with plant resistance. The ANAC001 protein provided by the inventioncan regulate and control the synthesis of salicylic acid in a plant so as to enhance plant disease resistance. The invention has a great value for the cultivation of disease resistant plants.

Description

technical field [0001] The invention relates to a plant resistance-related protein ANAC001 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 to...

Claims

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Application Information

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