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Application of rice ACCase mutant gene and protein thereof in plant herbicide resistance

A mutant, herbicide technology, applied in the field of plant genetic engineering and plant herbicide resistance breeding, to achieve the effect of significant resistance function

Pending Publication Date: 2021-02-26
JIANGSU ACADEMY OF AGRICULTURAL SCIENCES +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] At present, there is no research report that the ACCase site mutation in rice can be used to resist herbicides such as quizalofop-p-spergine and gaizalen

Method used

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  • Application of rice ACCase mutant gene and protein thereof in plant herbicide resistance
  • Application of rice ACCase mutant gene and protein thereof in plant herbicide resistance
  • Application of rice ACCase mutant gene and protein thereof in plant herbicide resistance

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0062] Embodiment 1 Rice M3 mutant is resistant to acetyl-CoA carboxylase herbicides (quinalofop-p-ethyl)

[0063] Use 0.5-1.0% (w / w) methanesulfonic acid in 150 kg of japonica conventional rice seed He Guang Si Miao (a gift from the Jiangsu Provincial Agricultural Germplasm Resources Protection and Utilization Platform, this is M0, soaked in water for 2 hours) 6 times Soak in ethyl ester (EMS) at room temperature for 6-9 hours, shake the seeds every 1 hour during this period; discard the EMS solution, soak the seeds 5 times in tap water, 5 minutes each time, then rinse the seeds with tap water overnight, and sow in the field the next day , and carry out conventional fertilizer and water management (this is M1). After the plants are mature, the seeds are mixed, dried, and stored for the winter. Sow in the field the following year. When the rice (this is M2) seedlings grow to the 3-4 leaf stage, spray 1mL quizalofop-p-ethyl / L water, and after 30 days, the normal green plants ...

Embodiment 2

[0066] Example 2: Obtaining the full-length gene of wild-type Accase in rice Zhennuo 19

[0067] Select the leaves of He Guang Si Miao wild-type plants, extract genomic DNA, design primers according to the Nipponbare ACCase gene (NCBI: XM_015783727) in NCBI to amplify, and use KOD DNApolymerase polymerase (purchased from Toyobo Company) to amplify the ACCase gene, the reaction The system is as follows:

[0068]

[0069] The PCR amplification reaction program adopts a two-step method, annealing and extension are combined together, and 68 degrees are used.

[0070] The program is as follows: pre-denaturation: 94°C for 3min; 35 cycles: denaturation at 94°C for 10sec; extension at 68°C for 10min; insulation: 72°C for 10min.

[0071] Take 2 μl of PCR products and detect by 1% agarose gel electrophoresis, after finding fragments of expected size ( image 3 ), the remaining PCR products were cleaned and recovered by a PCR cleaning kit (purchased from Axygen), cloned into the pMD...

Embodiment 3

[0076] Example 3: Mutation site analysis of rice mutant HGJK-3 resistant to acetyl-CoA carboxylase herbicides

[0077] The leaves of the obtained herbicide-resistant mutant rice plant HGJK-3 were selected and genomic DNA was extracted, and the genomic DNA of the wild-type rice He Guangsi seedlings and the rice mutant plant HGJK-3 were respectively used as templates for PCR amplification. Increase condition is the same as embodiment 2, amplification result is as image 3. At the same time, it was sent to Nanjing Yidao Biotechnology Co., Ltd. for sequencing. Comparing the sequencing results with the wild-type ACCase gene of Heguang Simiao, it was found that there was a mutation at one site on the ACCase gene, and the 6029th base was mutated from G to T, resulting in a change in the corresponding encoded amino acid sequence. The 2010th position is changed from tryptophan to leucine, that is, the nucleotide sequence of the ACCase gene of the herbicide-resistant mutant is shown i...

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Abstract

The invention discloses application of a rice ACCase mutant gene and protein thereof in plant herbicide resistance. The amino acid sequence of the rice ACCase mutant protein has the following mutation: the 2010 site and / or 2099<t>h site of the amino acid sequence corresponding to wild rice ACCase are mutated. The ACCase mutant gene is introduced into a rice genome sensitive to herbicide by a transgenic method and expressed, and a receptor plant can have resistance (tolerance) to acetyl coenzyme A carboxylase inhibitor herbicide. The ACCase mutant gene and the protein thereof can be widelyapplied to breeding of herbicide-resistant plants.

Description

technical field [0001] The invention relates to the application of rice ACCase mutant gene and its protein in plant herbicide resistance, and belongs to the fields of plant genetic engineering and plant herbicide resistance breeding. Background technique [0002] Acetyl-CoA carboxylase (ACCase) is the rate-limiting enzyme in the de novo fatty acid synthesis pathway in plants, and participates in the process of generating malonyl-CoA from acetyl-CoA as a precursor in the plastid. ACCase consists of three parts, namely biotin carboxyl carrier protein (BCCP), biotin carboxylase (BC) and carboxyltransferase (CT). Because ACCase plays a key role in the normal growth and development of plants, pesticide companies have developed a series of herbicides targeting ACCase, which have been promoted in the market so far. [0003] ACCase is a key enzyme in plant fatty acid biosynthesis. It catalyzes ATP-dependent acetyl-CoA carboxylation to induce a key metabolite in the biosynthesis of ...

Claims

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Application Information

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IPC IPC(8): C12N9/00C12N15/52C12N15/82A01H5/00A01H6/46C12Q1/6895G01N33/573A01G13/00
CPCC12N9/93C12Y604/01002C12N15/8274C12Q1/6895G01N33/573A01G13/00C12Q2600/156C12Q2600/13G01N2333/9015
Inventor 张保龙王金彦凌溪铁王凤斌
Owner JIANGSU ACADEMY OF AGRICULTURAL SCIENCES
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