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CN-122012595-A - Application of OsmiR159-GAMYBL2-RGA1 signaling pathway in regulation and control of rice drought tolerance

CN122012595ACN 122012595 ACN122012595 ACN 122012595ACN-122012595-A

Abstract

The application discloses application of an OsmiR159-GAMYBL2-RGA1 signal path in regulation and control of rice drought tolerance, in particular to an OsmiR159 signal transmission chain which is formed by directly targeting and inhibiting expression of OsGAMYBL transcription factors and a OsGAMYBL protein by combining a promoter of G protein alpha subunit encoding gene RGA1 and inhibiting transcription. Based on the application, a high-efficiency molecular target and an implementation technical scheme are provided for rice drought-enduring breeding.

Inventors

  • SHI ZHENYING
  • XIE LINGJUAN
  • WU SHUJUN
  • CAO LIMING
  • YAN YING
  • YANG HANG
  • WANG KAI
  • ZHANG LIXIA
  • YE JUNHUA
  • HU ZEJUN

Assignees

  • 上海市农业科学院

Dates

Publication Date
20260512
Application Date
20260310

Claims (10)

  1. Use of the osmir159-GAMYBL2-RGA1 signalling pathway for modulating drought tolerance in rice, characterised in that said modulation is achieved by: OsmiR159 negatively regulates OsGAMYBL gene expression, and relieves OsGAMYBL protein inhibition of RGA1 gene transcription, so that RGA1 gene-mediated drought-enduring negative regulation effect is reduced.
  2. 2. The use of the OsmiR 159-GAMYBL-RGA 1 signaling pathway of claim 1 for modulating drought tolerance in rice, wherein said modulation is to enhance survival of rice under drought stress.
  3. 3. The use of the OsmiR159-GAMYBL2-RGA1 signaling pathway of claim 2 for modulating drought tolerance in rice, wherein the OsGAMYBL protein binds to a MYB binding motif in the RGA1 promoter via its R2R3 domain, thereby inhibiting transcription of RGA 1.
  4. 4. A method for improving drought tolerance of rice, which is characterized in that the expression of OsmiR159 in the rice and/or the expression of RGA1 gene in the rice is/are down-regulated by the regulation mode of any one of claims 1-3, so that the survival rate of the rice under drought stress is improved.
  5. 5. The method of claim 4, wherein the down-regulating expression of OsmiR159 is achieved by inhibiting the activity or reducing the level of accumulation of OsmiR 159; And/or down-regulating RGA1 expression by inhibiting transcription or translation of RGA1 genes.
  6. 6. The method of claim 5, wherein the down-regulating expression of OsmiR159 is achieved by short tandem target mimetic STTM technology or gene editing technology; The STTM targeting sequence is 5'-UUCCACAGCUUUCUUGAACUG-3'; and/or said down-regulating RGA1 expression is effected by gene editing techniques or RNA interference techniques.
  7. 7. The method for breeding drought-enduring rice is characterized by comprising the following steps: a) Providing rice material with down-regulated OsmiR159 expression and/or providing rice material with down-regulated RGA1 gene expression; b) Hybridizing the rice material provided in step a) with a target rice variety; c) And screening the plant with the offspring having OsmiR159 low expression and/or RGA1 low expression and improved drought tolerance.
  8. 8. The method for breeding drought-tolerant rice according to claim 7, wherein the rice material whose expression is down-regulated by OsmiR159 is a material obtained by the method for improving drought tolerance of rice according to any one of claims 4 to 6.
  9. 9. The method for breeding drought-tolerant rice according to claim 7, wherein the rice material whose RGA1 gene expression is down-regulated is a material obtained by the method for improving drought tolerance of rice according to any one of claims 4 to 6.
  10. 10. A biomaterial for use in carrying out the method of any one of claims 4-6 and 7-9, the biomaterial being a recombinant vector comprising an STTM sequence targeting OsmiR159, or a CRISPR vector comprising an sgRNA sequence targeting RGA1 gene, or a host cell comprising the vector.

Description

Application of OsmiR159-GAMYBL2-RGA1 signaling pathway in regulation and control of rice drought tolerance Technical Field The invention belongs to the technical field of plant gene breeding, and particularly relates to application of an OsmiR159-GAMYBL2-RGA1 signal pathway in regulation and control of rice drought tolerance. Background Drought is one of the most important abiotic stresses restricting global agricultural production, and as climate change is aggravated, the frequency, duration and intensity of drought occurrence are continuously increased, which forms a serious threat to grain safety. The rice is used as staple food for nearly half of the population in the world, has large water demand for growth and development and is extremely sensitive to drought stress. Therefore, the molecular mechanism of rice drought tolerance is deeply analyzed, and new varieties of drought-tolerant water-saving rice are cultivated based on the molecular mechanism, so that the molecular mechanism has great strategic significance in guaranteeing the grain safety of China and the world. Plants evolved complex mechanisms to cope with drought stress, including but not limited to morphological adaptive changes, physiological and biochemical regulation, and stomatal movement regulation to reduce water transpiration, while at the molecular level drought stress induces extensive transcriptome, proteome and metabolome reprogramming involving the involvement of large numbers of transcription factors, kinases, and non-coding RNAs. MicroRNAs are a class of endogenous non-coding MicroRNAs of about 21-24 nucleotides in length, which play a key regulatory role in plant growth, development, metabolism and stress response by mediating cleavage or translational inhibition of target gene mRNA. In drought response, multiple mirnas have been demonstrated to be involved in regulation, e.g., miR396/GRF module, miR156/SPL module, miR393, etc., are reported to be associated with drought tolerance in crops such as arabidopsis, maize, etc. However, in rice, the specific functions and downstream regulatory networks of most drought-responsive miRNAs are still unclear, limiting their potential for use in genetic improvement of rice drought tolerance. Heterotrimeric G proteins are conserved eukaryotic signal transduction molecular switches. In rice, the G protein alpha subunit encoding gene RGA1 (also known as D1) has been reported to be involved in regulation of a variety of agronomic traits. Studies have shown that RGA1 loss-of-function mutant (d 1) exhibits some increase in drought tolerance during vegetative growth. However, the upstream regulatory factors of RGA1 and their precise molecular mechanisms in drought tolerance regulation have not been fully elucidated, and in particular their association with miRNA regulatory networks is not known. OsmiR159 is a highly conserved miRNA family in plants, and has the core function of targeted regulation of GAMYB transcription factors, and is involved in plant development, hormone response and stress adaptation. In arabidopsis, miR159 participates in an ABA signal path and influences stomatal closure, however, whether or not OsmiR159 functions in rice drought tolerance, how to function and whether or not the OsmiR159 is associated with a known drought tolerance related path (such as G protein signal) is not reported at present, and the technology blank in the field is provided. Based on the background, the application firstly illustrates a totally new drought-enduring regulation and control path for connecting miRNA, transcription factors and G protein signals through genetic, molecular and physiological experiments of a system, and develops a feasible rice drought-enduring molecular breeding strategy based on the discovery, thereby providing a new theoretical basis and technical means for coping with drought stress and cultivating green super rice. Disclosure of Invention In order to fill the research blank in the aspects of functions and mechanisms in the drought tolerance regulation of rice at the present stage, the application provides application of an OsmiR 159-GAMYBL-RGA 1 signal path in regulation and control of the drought tolerance of the rice, and the result has extremely high economic benefit in coping with drought stress and rice yield dimension. In a first aspect, the application provides an application of an OsmiR159-GAMYBL2-RGA1 signal pathway in regulation and control of rice drought tolerance, wherein the regulation and control are realized by the following modes: OsmiR159 negatively regulates OsGAMYBL gene expression, and relieves OsGAMYBL protein inhibition of RGA1 gene transcription, so that RGA1 gene-mediated drought-enduring negative regulation effect is reduced. Further, the regulation is to enhance the survival rate of rice under drought stress. Further, the OsGAMYBL protein binds to the MYB binding motif in the RGA1 promoter through its R2R3 domain, thereby inhibiting transcr