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KR-20260065988-A - Chimeric insecticidal proteins and their use

KR20260065988AKR 20260065988 AKR20260065988 AKR 20260065988AKR-20260065988-A

Abstract

The present invention relates to a chimeric insecticidal protein and its use, wherein the chimeric insecticidal protein comprises domains I and II of the Cry1Ac protein and domain III of the Cry1Ig protein, which are sequentially bound. The chimeric insecticidal protein possesses a potent body weight-inhibiting effect and lethal activity against various Lepidoptera pests. The chimeric insecticidal protein of the present invention not only broadens the insecticidal spectrum of the original protein but also enhances toxicity against pests, effectively slows down the development of pest resistance in nature, and has excellent prospects for application in agricultural production.

Inventors

  • 왕 친양
  • 팡 제
  • 셰 샹팅
  • 장 량웨이
  • 자 샤오웨이

Assignees

  • 베이징 다베이농 바이오테크놀로지 컴퍼니 리미티드

Dates

Publication Date
20260512
Application Date
20241031

Claims (16)

  1. As a chimeric insecticidal protein, The above chimeric insecticidal protein comprises domains I and II of the Cry1Ac protein and domain III of the Cry1Ig protein, which are sequentially bound. Chimeric insecticidal protein.
  2. In paragraph 1, The above chimeric insecticidal protein comprises domains I and II of the Cry1Ac protein, domain III of the Cry1Ig protein, the protoxin tail of the Cry1Ac protein, or a fragment thereof, which are sequentially bound. Chimeric insecticidal protein.
  3. In paragraph 1 or 2, The above chimeric insecticidal protein comprises the amino acid sequence shown in SEQ ID NO. 1 or SEQ ID NO. 2, Chimeric insecticidal protein.
  4. Encoding a chimeric insecticidal protein according to any one of paragraphs 1 to 3, Nucleic acid molecule.
  5. In paragraph 4, The nucleic acid molecule comprises the sequence shown in SEQ ID NO. 3, SEQ ID NO. 4, SEQ ID NO. 5, or SEQ ID NO. 6, Nucleic acid molecule.
  6. A nucleic acid molecule comprising either paragraph 4 or 5, Expression cassette.
  7. A nucleic acid molecule according to either paragraph 4 or 5, or comprising an expression cassette according to paragraph 6, Expression vector.
  8. A chimeric insecticidal protein according to any one of claims 1 to 3, a nucleic acid molecule according to any one of claim 4 or 5, an expression cassette according to claim 6, or an expression vector according to claim 7, Host cell.
  9. As a plant or part of a plant, The above plant or part of the plant comprises a chimeric insecticidal protein according to any one of claims 1 to 3, a nucleic acid molecule according to any one of claim 4 or 5, an expression cassette according to claim 6, or an expression vector according to claim 7, Plant or part of a plant.
  10. A method for producing a transgenic plant comprising the following steps: 1) A step of transforming a nucleic acid molecule according to claim 4 or 5, an expression cassette according to claim 6, or an expression vector according to claim 7 into a plant cell; 2) a step of selecting a plant cell comprising the above nucleic acid molecule, expression cassette, or expression vector; and 3) Step of regenerating selected plant cells into plants.
  11. As an insecticidal composition, The above insecticidal composition comprises a chimeric insecticidal protein according to any one of claims 1 to 3, Insecticide composition.
  12. As a method of controlling Lepidoptera pests, The above method comprises the steps of: contacting a Lepidoptera pest with a chimeric insecticidal protein according to any one of claims 1 to 3, a host cell according to claim 8, a plant or part of a plant according to claim 9, or an insecticidal composition according to claim 11; or introducing a nucleic acid molecule according to any one of claims 4 or 5, an expression cassette according to claim 6, or an expression vector according to claim 7 into a plant, and contacting the Lepidoptera pest with said plant. Methods for controlling Lepidoptera pests.
  13. In Paragraph 12, The above Lepidoptera pests include Noctuidae pests or Pyralidae pests; Preferably, Lepidoptera pests include Spodoptera frugiperda , Spodoptera litura , Spodoptera exigua , Argyrogramma agnata , Mythimna separata , Helicoverpa armigera , and/or Ostrinia furnacalis , Methods for controlling Lepidoptera pests.
  14. Use of a chimeric insecticidal protein according to any one of claims 1 to 3, a nucleic acid molecule according to any one of claim 4 or 5, an expression cassette according to claim 6, an expression vector according to claim 7, or an insecticidal composition according to claim 11 for controlling Lepidoptera pests.
  15. In Paragraph 14, The above use comprises the step of contacting the Lepidoptera pest with a chimeric insecticidal protein according to any one of claims 1 to 3 or an insecticidal composition according to claim 11 to control the Lepidoptera pest, or introducing a nucleic acid molecule according to any one of claim 4 or 5, an expression cassette according to claim 6, or an expression vector according to claim 7 into a plant to control the Lepidoptera pest, and bringing the Lepidoptera pest into contact with the plant. Preferably, the Lepidoptera pests include Noctuidae pests or Pyralidae pests; More preferably, the Lepidoptera pest comprises Spodoptera frugiperda , Spodoptera litura , Spodoptera exigua , Argyrogramma agnata , Mythimna separata , Helicoverpa armigera , and/or Ostrinia furnacalis , use.
  16. As a product, The above product is a product obtained from a plant or part of a plant according to claim 9, or from a genetically modified plant obtained by a method for producing a genetically modified plant according to claim 10, wherein the product is a grain, starch, oil, syrup, flour, crushed meal, cereal, or protein; Preferably, the product comprises a chimeric insecticidal protein according to any one of claims 1 to 3, or a nucleic acid molecule according to any one of claims 4 or 5, product.

Description

Chimeric insecticidal proteins and their use The present invention relates to a chimeric insecticidal protein, a nucleic acid molecule encoding the protein, and a method and use thereof for controlling Lepidoptera pests. Currently, agricultural production faces biological stress (such as diseases and pests) and abiotic stress (such as drought, cold, and salinity damage), which weaken crop growth and reduce yields, posing a serious threat to global food security. Among these, pests are one of the major biological stressors affecting agricultural and forestry productivity. As environmental problems caused by the use of chemical pesticides for pest control become increasingly severe, public interest in the use of biological pesticides is gradually rising. Bacillus thuringiensis ( Bt) is a Gram-positive bacterium widely distributed in nature and is also an insect pathogen. The most distinct feature distinguishing Bt bacteria from other Bacillus species is the formation of crystal proteins along with spore formation during the later stages of growth, which are generally referred to as spore crystals (including Cry and Cyt proteins). These proteins are major or decisive factors in the development of insect pathogenicity. A large number of studies have reported that various Bt proteins (Cry1Ab, Cry1Ac, Cry1F, Cry2Ab, and Cry3Bb) have insecticidal activity against Lepidoptera , Coleoptera , Diptera , Hymenoptera , and Homoptera insects. The commercial cultivation of Bt transgenic insect-resistant crops produced by plant genetic engineering has also become one of the major drivers significantly improving agricultural productivity. These transgenic insect-resistant plants not only effectively control the occurrence and damage caused by target pests but also provide important assurances for food and ecological security by reducing the application of chemical pesticides. However, with the popularization and application of genetically modified crops, insects will evolve resistance to the Bt proteins expressed in these plants under continuous selective pressure. If this resistance cannot be effectively controlled, it will limit the commercial value of genetically modified plants containing Bt proteins. Meanwhile, some Bt proteins possess potent cross-lethal activity against non-target organisms as well. All of these issues may limit the further application of Bt proteins. The rational design of functionally enhanced Bt chimeric proteins (also referred to as fusion proteins) based on Bt protein domains is one method to address the aforementioned problems. Using this method, Bt chimeric proteins can be obtained by introducing or replacing domains of heterologous Bt proteins or other types of insecticidal proteins into the Bt protein, or by recombining different domains of the Bt protein, thereby possessing potent insecticidal activity, a broader insecticidal spectrum, and even the ability to offset insecticide resistance in target pests. Currently, related research is primarily focused on the recombination of Bt Cry protein domains that possess clear structures and functions. Even if extensive analysis, design, screening, and validation are performed, it is not possible to reliably construct chimeric insecticidal proteins that exhibit enhanced insecticidal activity compared to parent proteins. Therefore, methods to design novel chimeric proteins with improved insecticidal activity and a broader insecticidal spectrum to provide more selective insecticidal proteins suitable for use in agricultural production remain a key research challenge within the industry. Considering the disadvantages of the prior art, the present invention provides a chimeric insecticidal protein capable of simultaneously being toxic to various Lepidoptera pests. The chimeric insecticidal protein has a strong body weight inhibiting effect and lethal activity against Lepidoptera pests including Spodoptera frugiperda , Helicoberta armigera , Argyrogramma agnata , Spodoptera litura, Mythimna separata , and Ostrinia furnacalis . Plants into which the coding gene for these proteins has been introduced exhibit potent insect resistance activity, particularly against Spodoptera frugiperda and Helicoberta armigera , with lethality rates of up to 61% and 88%, respectively. The protein provided by the present invention is a novel chimeric insecticidal protein. Compared to the insect resistance protein before modification, the chimeric insecticidal protein provided by the present invention has a broader insecticidal spectrum and higher insecticidal activity, can effectively delay the development of insect resistance in nature, and has excellent prospects for application in agricultural production. In a first aspect, the present invention provides a chimeric insecticidal protein, wherein the chimeric insecticidal protein comprises a first domain (domain I) and a second domain (domain II) of a Cry1Ac protein and a third domain (domain III) of a Cry1Ig protein that a