Rice herbicide-resistant als mutant protein, nucleic acid and application thereof

A resistant protein and mutant technology, applied in the field of plant protein and plant herbicide resistance, can solve the problems that it is difficult to predict the herbicide resistance of ALS protein in advance, and the mechanism of herbicide action is not determined

Active Publication Date: 2019-04-26
JIANGSU ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] At present, the mechanism of action of ALS inhibitor herbicides has not yet been determined, and it is difficult to predict in advance whether mutations at other amino acid sites of the ALS protein will cause herbicide resistance. New herbicide-resistance locus in ALS protein just might be discovered by luck

Method used

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  • Rice herbicide-resistant als mutant protein, nucleic acid and application thereof
  • Rice herbicide-resistant als mutant protein, nucleic acid and application thereof
  • Rice herbicide-resistant als mutant protein, nucleic acid and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0060] Example 1: Obtaining the full-length ALS gene of Huazhan wild-type rice

[0061] The genomic DNA of Huazhan, an indica-type conventional rice seed, was extracted by conventional methods. The indica-type conventional rice seed Huazhan was donated by Yangzhou Academy of Agricultural Sciences. For information about the relative parents of Huazhan, please refer to China Rice Research Institute http: / / www.ricedata.cn / variety / varis / 607962.htm? 607962 According to the Nipponbare ALS gene (NCBI: XM_015770973) in NCBI, primers were designed for amplification, and Takara PrimerSTAR Max DNA Polymerase polymerase (purchased from Takara Company) was used to amplify the ALS gene. The reaction system was as follows:

[0062] 2×PrimerSTAR Max Premix 10.0μl

[0063] 10 μM forward primer 1.0 μl

[0064] 10 μM reverse primer 1.0 μl

[0065] 20-30ng / μL Huazhan wild-type rice genomic DNA 1.0μl

[0066] Add sterile water to a total volume of 20 μl

[0067] The PCR amplification reac...

Embodiment 2

[0071] Example 2: The process of obtaining rice mutants resistant to imidazolinone herbicides (Bailongtong)

[0072] For information about the relative parents of Huazhan, an indica-type conventional rice seed (gifted by Yangzhou Academy of Agricultural Sciences), see China Rice Research Institute http: / / www.ricedata.cn / variety / varis / 607962.htm? 607962 ,

[0073] (This is M0, soaked in water for 2 hours) 150kg was divided into 6 times and soaked with 0.5-1.0% (w / w) ethyl methanesulfonate (EMS) at room temperature for 6-9 hours, during which the seeds were shaken every 1 hour; Remove the EMS solution, turn and soak the seeds 5 times in tap water, 5 minutes each time, then rinse the seeds with tap water overnight, sow in the field the next day, and perform 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 rice (this is M2) seedlings grow to ...

Embodiment 3

[0074] Example 3: Analysis of mutation sites in rice mutants resistant to imidazolinone herbicides

[0075] Genomic DNA was extracted from the leaves of the Huazhan herbicide-resistant rice mutant-1 plant and the Huazhan herbicide-resistant rice mutant-2 plant obtained in Example 2 above, and sent to Nanjing Yidao Biotechnology Co., Ltd. for genome sequencing. Comparing the sequencing results with the Huazhan wild-type ALS gene, it was found that both of the two resistant rice mutants mutated at position 627 of the ALS gene, and at position 1879 of the ALS gene sequence in the Huazhan herbicide-resistant rice mutant-1 The nucleotide changes from A to C, and the 1880th nucleotide from G to T, resulting in the 627th position of the corresponding encoded amino acid sequence changing from serine to leucine, that is, Huazhan The nucleotide sequence of the ALS gene of the herbicide-resistant rice mutant-1 is shown in SEQ ID NO.1, and the amino acid sequence of the ALS protein encode...

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Abstract

The invention discloses a rice herbicide resistant ALS mutant protein. The amino acid sequence of the ALS mutant protein has any one of the following mutations: serine which corresponds to an amino acid at a 627th position of rice ALS is changed into leucine, or serine which corresponds to an amino acid at a 627th position of rice ALS is changed into cysteine. The invention further discloses a nucleic acid and application of an expression cassette, a recombinant vector or cells in in the aspect of a weed resisting and removing agent for plants. A result of an experiment of spraying an ALS inhibitors herbicide 'imazameth' in a field shows that after 3.3 mL of imazameth / L water is applied to rice 3-4-leaf seedlings which separately contains two ice herbicide resistant ALS mutant proteins ofthe invention, plants still grow and develop normally and are firm, but 30 days after 1 mL imazameth / 3L water is applied to wild rice 3-4-leaf seedling, the whole plants are dead.

Description

technical field [0001] The invention belongs to the field of plant protein and plant herbicide resistance, and relates to rice ALS mutant protein, gene and application thereof that make plants have herbicide resistance; specifically, the present invention relates to rice acetolactate synthase (ALS) mutant protein , the protein can endow plants, especially rice, with the characteristics of resistance to acetolactate synthase inhibitor herbicides. The invention discloses the sequence of the protein and their application in the field of plant herbicide resistance. Background technique [0002] Weeds are unfavorable factors restricting the stable and high yield of agricultural production. Compared with the traditional methods of relying on cultivation measures, manual weeding and mechanical weeding, the use of chemical herbicides is an efficient, simple and economical method of controlling weeds. [0003] At present, the acetolactate synthase (ALS) inhibitor chlorsulfuron has ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N9/88C12N15/60C12Q1/6895A01H5/00A01H6/46A01H6/82A01H6/60A01H6/20A01N25/32A01N43/50A01P13/00
CPCA01N25/32A01N43/50C12N9/88C12N15/8274C12Q1/6895C12Q2600/13C12Y401/03
Inventor 张保龙王金彦凌溪铁陈天子
Owner JIANGSU ACAD OF AGRI SCI
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