Search

CN-121975823-A - Herbicide-resistant rice OsACC gene mutant and application thereof

CN121975823ACN 121975823 ACN121975823 ACN 121975823ACN-121975823-A

Abstract

The invention relates to herbicide-resistant rice OsACC gene mutant and application thereof. The rice OsACC gene mutant is a deletion mutant, the sequence of nucleic acid before deletion mutation is shown as SEQ ID No. 1, the length of the sequence of deleted nucleic acid is 33 to 97 bp, A in the ATG of OsACC gene initiation codon is 0, the deleted nucleic acid occurs in 305 to 209 sites upstream of the ATG of initiation codon and at least in 251 to 219 sites upstream of the ATG of initiation codon, and the deletion mutant is excluded that the length of the sequence of deleted nucleic acid is 53 to 97 bp, the deleted nucleic acid occurs in 305 to 209 sites upstream of the ATG of initiation codon and at least in 271 to 219 sites upstream of the ATG of initiation codon. The mutant can enable the rice to obtain three resistances of high-efficiency haloxyfop-methyl, haloxyfop-methyl and clethodim.

Inventors

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

Assignees

  • 中国农业科学院植物保护研究所

Dates

Publication Date
20260505
Application Date
20260403

Claims (10)

  1. 1. A rice OsACC gene mutant, a rice OsACC gene mutant, wherein the rice OsACC gene mutant is a deletion mutant; wherein the deletion mutation is in the promoter of rice OsACC gene, the sequence of the nucleic acid before deletion mutation is shown as SEQ ID No. 1, the length of the deleted nucleic acid is 33 bp to 97 bp, A in ATG of OsACC gene initiation codon is 0, the deleted nucleic acid is in 305 to 209 sites upstream of ATG initiation codon and at least deletes the 251 to 219 bases upstream of ATG initiation codon, and The mutants were deleted whose sequence was 53 bp to 97 bp in length, whose A was 0 in the ATG of the OsACC gene start codon, which occurred in positions 305 to 209 upstream of the ATG of the start codon, and whose bases at least 271 to 219 upstream of the ATG of the start codon were deleted.
  2. 2. The rice OsACC gene mutant according to claim 1, wherein the length of the sequence of the deleted nucleic acid is 33 bp to 92 bp, wherein the a in the OsACC gene start codon ATG is 0, the deleted nucleic acid occurs within 300 to 209 sites upstream of the start codon ATG, and at least the base located 251 to 219 sites upstream of the start codon ATG is deleted.
  3. 3. The rice OsACC gene mutant according to claim 1, wherein the length of the sequence of the deleted nucleic acid is 33 bp to 73 bp, wherein the a in the OsACC gene start codon ATG is 0, the deleted nucleic acid occurs in positions 281 to 209 upstream of the start codon ATG, and at least the base located 251 to 219 upstream of the start codon ATG is deleted.
  4. 4. The rice OsACC gene mutant according to claim 1, wherein the length of the sequence of the deleted nucleic acid is 33 bp to 63 bp, wherein the a in the OsACC gene start codon ATG is 0, the deleted nucleic acid occurs within the 271 to 209 positions upstream of the start codon ATG, and at least the 251 to 219 bases upstream of the start codon ATG are deleted.
  5. 5. The rice OsACC gene mutant according to claim 1, wherein the length of the sequence of the deleted nucleic acid is 33 bp to 54 bp, wherein the a in the OsACC gene start codon ATG is 0, the deleted nucleic acid occurs in the 262 to 209 positions upstream of the start codon ATG, and at least the 251 to 219 bases upstream of the start codon ATG are deleted.
  6. 6. A rice OsACC gene mutant according to claim 5, wherein the deleted nucleic acid has a sequence length of 33 bp and the A in the ATG of the OsACC gene is 0, the deleted nucleic acid occurs at 251 to 219 positions upstream of the ATG of the initiation codon, and/or The deleted nucleic acid was 54 bp in length and occurred at positions 262 to 209 upstream of the initiation codon ATG with A in the initiation codon ATG of the OsACC gene being 0.
  7. 7. Use of a rice OsACC gene mutant according to any one of claims 1 to 6 in a herbicide against ACCase inhibitors in rice of the type south japonica 46 or japan.
  8. 8. The use according to claim 7, wherein the ACCase inhibitor herbicide is at least one of haloxyfop-methyl, haloxyfop-methyl and clethodim.
  9. 9. A method for obtaining the capability of rice to resist ACCase inhibitor herbicides, which is realized by deleting partial base in OsACC gene promoter in rice genome by gene editing or homologous recombination to obtain rice OsACC gene mutant according to any one of claims 1-6, wherein the rice variety is Nanjing 46 or Nippon Temminck.
  10. 10. The method according to claim 9, 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 Nos. 7 or 8, and the target sequence is positioned at 34 th to 56 th positions in the sequence shown as SEQ ID Nos. 7 or 8; 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.

Description

Herbicide-resistant rice OsACC gene mutant and application thereof Technical Field The invention relates to the field of nucleic acid, in particular to herbicide-resistant rice OsACC gene mutant 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 One of the present invention provides a rice OsACC gene mutant, which is a deletion mutant, wherein the deletion mutation occurs in the promoter of the rice OsACC gene, the sequence of the nucleic acid before the deletion mutation is shown as SEQ ID No.1, the length of the deleted nucleic acid is 33 bp to 97 bp, A in the start codon ATG of the OsACC gene is 0, the deleted nucleic acid occurs in the 305 to 209 sites upstream of the start codon ATG (the sequence of which is shown as SEQ ID No. 11) and at least the 251 to 219 bases upstream of the start codon ATG (the sequence of which is shown as SEQ ID No. 12) is deleted, and the deletion mutant is excluded, the length of the sequence of the deleted nucleic acid is 53 bp to 97 bp, A in the start codon ATG of the OsACC gene is 0, the deleted nucleic acid occurs in the 305 to 209 sites upstream of the start codon ATG (the sequence of which is shown as SEQ ID No. 11) and at least the 271 to 219 bases upstream of the start codon ATG (the sequence of which is shown as SEQ ID No. 13) is deleted. That is, the deletion sequence may be SEQ ID No.11 containing SEQ ID No. 12 at the longest, or any one of SEQ ID No.11 containing SEQ ID No. 12 but excluding any one of SEQ ID No.11 containing SEQ ID No. 13, or any one of SEQ ID No.11 containing a combination of SEQ ID No. 12 and any other sequence of SEQ ID No.11 but excluding any one of SEQ ID No.11 containing a combination of SEQ ID No. 13 and any other sequence of SEQ ID No. 11. In a specific embodiment, the deleted nucleic acid has a sequence length of 33 bp to 92 bp, wherein at position 0 a in the OsACC gene start codon ATG, the deleted nucleic acid occurs within positions 300 to 209 upstream of the start codon ATG, and at least the bases 251 to 219 upstream of the start codon ATG are deleted. Among these, it is desirable to exclude mutants in which the length of the sequence of the deleted nucleic acid is 53 bp to 92 bp, the A in the initiation codon ATG of OsACC gene is 0, the deleted nucleic acid occurs in the 300 to 209 positions upstream of the initiation codon ATG, and at least the 271 to 219 bases upstream of the initiation codon ATG are deleted. In a specific embodiment, the deleted nucleic acid has a sequence length of 33 bp to 73: 73 bp, wherein the A in the ATG at the start codon of the OsACC gene is 0, the deleted nucleic acid occurs in positions 281 to 209 upstream of the ATG at least deleting the bases at positions 251 to 219 upstream of the ATG at the start codon. Among these, it is desirable to exclude mutants in which the length of the sequence of the deleted nucleic acid is 53 bp to 73: 73 bp, the A in the initiation codon ATG of OsACC gene is 0, the deleted nucleic acid occurs in positions 281 to 209 upstream of the initiation codon ATG, and at least the base at positions 271 to 219 upstream of the initiation codon ATG is deleted. In a specific embodiment, the deleted nucleic acid has a sequence length of 33 bp to 63 bp, wherein at position 0 a in the OsACC gene start codon ATG, the deleted nucleic acid occurs within positions 271 to 209 upstream of the start codon ATG, and at least the bases 251 to 219 upstream of the start codon ATG are deleted. Among these, it is desirable to exclude mutants in which the length of the sequence of the deleted nucleic acid is 53 bp to 63 bp, the A in the initiation