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CN-121991978-A - Rice OsACC gene mutant for resisting haloxyfop-R-methyl or clethodim and application thereof

CN121991978ACN 121991978 ACN121991978 ACN 121991978ACN-121991978-A

Abstract

The invention relates to a rice OsACC gene mutant for resisting haloxyfop-R-methyl or clethodim and application thereof. The rice OsACC gene mutant is a deletion mutant or a deletion-combination insertion mutant, wherein the deletion mutation is generated in a promoter of a rice OsACC gene, the sequence of nucleic acid before the deletion mutation is shown as SEQ ID No.1, the length of the sequence of deleted nucleic acid is 16 bp, A in an ATG of OsACC genes is 0, the deleted nucleic acid is generated in 455-440 positions upstream of the ATG of the OsACC genes, A in the ATG of OsACC genes is 0, and the inserted nucleic acid is generated in 455-440 positions upstream of the ATG of the initiation genes. The rice OsACC gene mutant can enable rice to obtain two resistances to herbicide haloxyfop-R-methyl and clethodim.

Inventors

  • YAN FANG
  • Yan Junlian
  • ZHOU HUANBIN
  • REN BIN

Assignees

  • 中国农业科学院植物保护研究所

Dates

Publication Date
20260508
Application Date
20260403

Claims (8)

  1. 1. A rice OsACC gene mutant is a deletion mutant, wherein the deletion mutation is generated in a promoter of a rice OsACC gene, the sequence of nucleic acid before the deletion mutation is shown as SEQ ID No. 1, the length of the sequence of deleted nucleic acid is 16 bp, wherein A in an ATG of a OsACC gene start codon is 0, and the deleted nucleic acid is generated in 455-440 positions upstream of the ATG of the start codon.
  2. 2. A rice OsACC gene mutant is a deletion-combining insertion mutant, wherein the deletion-combining insertion mutant is generated in a promoter of a rice OsACC gene, a sequence of nucleic acid before deletion-combining insertion mutation is shown as SEQ ID No. 1, the length of the sequence of deleted nucleic acid is 16 bp, the length of the sequence of inserted nucleic acid is at least 1 to 6 bp, A in an ATG of OsACC gene is 0, the deleted nucleic acid is generated at 455 to 440 positions upstream of the ATG of the initiation codon, A in an ATG of OsACC gene is 0, and the inserted nucleic acid is generated at 455 to 440 positions upstream of the ATG of the initiation codon.
  3. 3. The rice OsACC gene mutant according to claim 2, wherein the inserted nucleic acid has a sequence of any one of paragraphs 1 to 6 bp of TAATTA.
  4. 4. Use of a rice OsACC gene mutant according to any one of claims 1 to 3 in a herbicide against ACCase inhibitors in rice of the variety south japonica 46 or japan.
  5. 5. The use according to claim 4, wherein the ACCase inhibitor herbicide is haloxyfop-methyl and/or clethodim.
  6. 6. A method for obtaining a rice plant resistant to ACCase inhibitor herbicides by deleting a part of the base in OsACC gene promoter in the genome of the rice plant by gene editing or homologous recombination, and optionally inserting a part of the base into the OsACC gene promoter to obtain a rice plant OsACC gene mutant according to any one of claims 1 to 3, the rice plant being of the variety south japonica 46 or japan.
  7. 7. The method according to claim 6, characterized in that it comprises the steps of: 1) Obtaining a pHZLib-Cas12i3 vector or pHZ and a pUbi-IEE-Cas12i3 vector, wherein the pHZLib-Cas12i3 vector is constructed by replacing a DR-crRNA-BsaI-BsaI-DR element in pHZ33 with a suicide gene ccdB to obtain a pHZ-ccdB vector, integrating pHZ-ccdB and pUbi-IEE-Cas12i3 into one vector to obtain a pHZLib-Cas12i3 vector; 2) Obtaining a crRNA sequence or a target sequence for gene editing, wherein the crRNA sequence is shown as SEQ ID No. 6, and the target sequence is positioned at 34 th to 56 th positions in the sequence shown as SEQ ID No. 6; 3) Replacing the ccdB gene on the pHZLib-Cas12i3 vector with the crRNA, thereby cloning the crRNA sequence into the pHZLib-Cas12i3 vector to obtain pHZLib-Cas12i3-OsACCcrRNA, or cloning the target sequence into the pHZ33 vector to obtain pHZ-OsACCSpacer vector, and integrating the pHZ-OsACCSpacer and the pUbi-IEE-Cas12i3 vector into one vector to obtain pUbi-IEE-Cas12i3-HZ33-OsACCSpacer; 4) pHZLib-Cas12i3-OsACCcrRNA or pUbi-IEE-Cas12i3-HZ33-OsACCSpacer are transformed into agrobacterium and infect rice callus, and rice strains resistant to ACCase inhibitor herbicides are screened.
  8. 8. The method of claim 6 or 7, wherein the ACCase inhibitor herbicide is haloxyfop-methyl and/or clethodim.

Description

Rice OsACC gene mutant for resisting haloxyfop-R-methyl or clethodim and application thereof Technical Field The invention relates to the field of nucleic acid, in particular to a rice OsACC gene mutant for resisting haloxyfop-R-methyl or clethodim and application thereof. Background The field weeds are one of main biological disasters in agricultural production, and seriously affect the high-quality development of the rice industry. Among weed control strategies, the combined use of chemical control and herbicide resistant varieties has excellent performance. The Acetyl-CoA carboxylase (ACCase) inhibitor herbicide is mainly used for controlling grassy weeds, and has the characteristics of high efficiency, low toxicity, safety to succeeding crops and the like. At present, the technical means of cultivating ACCase inhibitor herbicide-resistant crops mainly comprise gene editing and the like, wherein the technical means comprise that mutation is carried out on a gene coding region of target protein ACC of the crop and the mutation of coded amino acid is generated, so that the binding capacity of the target protein after the mutation of the amino acid to herbicide is reduced, and the influence on normal vital activities of plants and the generation of herbicide resistance are avoided. However, such protein mutations often cause changes in the enzymatic activity of ACC itself and defects in growth and development, which limit its use in production. Furthermore, frequent use of a single herbicide accelerates the evolution of weed resistance and creates phytotoxicity problems. These problems greatly limit weed control in rice production. Disclosure of Invention The invention provides a rice OsACC gene mutant, which is a deletion mutant, wherein the deletion mutation is generated in a promoter of a rice OsACC gene, the sequence of nucleic acid before the deletion mutation is shown as SEQ ID No. 1, the length of the sequence of deleted nucleic acid is 16 bp, wherein A in an ATG of a OsACC gene start codon is 0, and the deleted nucleic acid is generated in 455-440 positions upstream of the ATG of the start codon. The invention also provides a rice OsACC gene mutant, which is a deletion-combining insertion mutant, wherein the deletion-combining insertion mutant is generated in a promoter of a rice OsACC gene, a sequence of nucleic acid before the deletion-combining insertion mutation is shown as SEQ ID No. 1, the length of the sequence of deleted nucleic acid is 16 bp, the length of the sequence of inserted nucleic acid is at least 1 to 6 bp, A in an initiating codon ATG of OsACC gene is 0, the deleted nucleic acid is generated in 455 to 440 positions upstream of the initiating codon ATG, A in an initiating codon ATG of OsACC gene is 0, and the inserted nucleic acid is generated in 455 to 440 positions upstream of the initiating codon ATG. In a specific embodiment, the inserted nucleic acid has a sequence of any one of paragraphs 1 to 6 bp of TAATTA. The invention also provides application of the rice OsACC gene mutant according to any one of the first and second aspects of the invention in the application of the rice ACCase inhibitor herbicide, wherein the rice is of the variety Nanjing 46 or Nippon Temminck. In a specific embodiment, the ACCase inhibitor herbicide is haloxyfop-methyl and/or clethodim. The fourth invention provides a method for obtaining the capability of resisting ACCase inhibitor herbicides of rice, which is realized by deleting partial bases in OsACC gene promoters in rice genome through gene editing or homologous recombination and optionally inserting partial bases into OsACC gene promoters to obtain the rice OsACC gene mutant according to any one of the first invention and the second invention, wherein the rice variety is south japonica 46 or Japanese sunny. In one embodiment, the method comprises the steps of: 1) Obtaining a pHZLib-Cas12i3 vector or pHZ and a pUbi-IEE-Cas12i3 vector, wherein the pHZLib-Cas12i3 vector is constructed by replacing a DR-crRNA-BsaI-BsaI-DR element in pHZ33 with a suicide gene ccdB to obtain a pHZ-ccdB vector, integrating pHZ-ccdB and pUbi-IEE-Cas12i3 into one vector to obtain a pHZLib-Cas12i3 vector; 2) Obtaining a crRNA sequence or a target sequence for gene editing, wherein the crRNA sequence is shown as SEQ ID No. 6, and the target sequence is positioned at 34 th to 56 th positions in the sequence shown as SEQ ID No. 6; 3) Replacing the ccdB gene on the pHZLib-Cas12i3 vector with the crRNA, thereby cloning the crRNA sequence into the pHZLib-Cas12i3 vector to obtain pHZLib-Cas12i3-OsACCcrRNA, or cloning the target sequence into the pHZ33 vector to obtain pHZ-OsACCSpacer vector, and integrating the pHZ-OsACCSpacer and the pUbi-IEE-Cas12i3 vector into one vector to obtain pUbi-IEE-Cas12i3-HZ33-OsACCSpacer; 4) pHZLib-Cas12i3-OsACCcrRNA or pUbi-IEE-Cas12i3-HZ33-OsACCSpacer are transformed into agrobacterium and infect rice callus, and rice strains resistant to ACCase