Wheat dwarf creeping alleles and their encoded proteins and applications

Abstract

The present invention discloses a wheat dwarf creeping allele and its encoding protein and application, belonging to the field of genetic engineering technology. Specifically disclosed are protein TaRHT-A1p (SEQ ID NO: 1) and TaRHT‑A1p Gene (SEQ ID NO: 2) and its application. Experiments have shown that increasing the content and / or activity of the protein TaRHT‑A1p in target plants can significantly reduce plant height, increase the plant's tillering angle, weaken the plant's ability to perceive gravity, and / or weaken the plant's response to gravity. The present invention successfully created dwarf wheat with significantly reduced plant height and significantly increased tillering angle. Dwarf wheat is less likely to fall over and has greater adaptability. The increased tillering angle can improve wheat's photosynthetic utilization efficiency, enhance field competitiveness, reduce pests and diseases, and increase yield. The present invention provides a new approach for the cultivation of new high-yield wheat varieties and provides a good genetic resource for wheat breeding.

Description

Technical Field

[0001] The invention belongs to the technical field of genetic engineering, and in particular relates to a wheat dwarf creeping allele and its encoded protein and application. Background Art

[0002] Wheat is an important food crop in the world. 20% of the calories and 25% of the protein consumed by the global population come from wheat. Wheat is also an important source of trace elements. Increasing wheat production is of vital importance to ensuring food security.

[0003] Plant height is a major component of wheat plant architecture and has a significant impact on wheat yield and other traits. Plant height closely influences lodging resistance and yield potential, making it one of the most important agronomic traits in genetic breeding. Lodging is a major factor affecting yield and quality in wheat production. Researchers have estimated that lodging-related grain yield losses can reach as high as 25% to 50% (Dunca J. Mechanical properties of cereal stem [J]. Research in Agricultural Engineering - UZPI (Czech Republic), 2008). Therefore, improving lodging resistance in wheat varieties by regulating plant height has long been a key goal in wheat breeding.

[0004] Plant height is negatively correlated with lodging resistance within a certain range. Therefore, dwarfing wheat plants can improve lodging resistance and achieve stable yields. However, excessively low plant height and short plant form can result in overly dense leaves, hindering ventilation and light transmission, thereby reducing yield. Therefore, dwarfing should be done in moderation during breeding. Research has shown that plant characteristics such as stem toughness, along with stem height, can influence lodging resistance, leading to some taller varieties being less prone to lodging than shorter varieties. However, with increasing water and fertilizer usage and improved cultivation techniques in wheat production, the lodging problem in tall wheat has intensified, increasing harvesting difficulties and reducing yield and processing quality. Dwarf wheat plants, because their stems are tougher than taller plants, possess greater lodging resistance. Since the Green Revolution, the use of dwarfing genes has been widely used and valued in breeding. The discovery and utilization of new dwarfing gene loci will be a long-term research hotspot in wheat.

[0005] Tiller angle has a profound impact on crop yield and has long attracted the attention of plant breeders and botanists. It is one of the key traits urgently in need of in-depth research and optimization in modern agricultural production. The formation of plant tiller angle is closely related to gravitropic response. Gravitropic response refers to the phenomenon in which plants, in response to gravity, readjust their growth direction to maintain the optimal angle between their organs and gravity. In other words, plants can sense gravity and adjust their growth direction. How plants perceive (or sense) gravity has always been a difficult and hot topic in plant science research. By regulating plant gravity perception, it is possible to further control the plant's gravitropic response (gravitropic response) and tiller angle. When gravity perception is weakened, the gravitropic response is weakened, resulting in a larger tiller angle. A larger tiller angle improves crop resistance to disease and competition for environmental resources, but it also reduces yield per unit area. Extremely compact crops, with smaller tiller angles, have less efficient light utilization, reduced resistance to pests and diseases, and are not conducive to increasing crop yield. Elucidating the molecular mechanisms regulating crop tiller angle will allow the breeding of more high-yield crop varieties suitable for cultivation in diverse environments. Wheat's prostrate trait is a wild-type trait, observed to varying degrees in diploid progenitors, certain local varieties, and key breeding parents (such as the Zhoumai series). Prostrate trait influences wheat plant architecture and tolerance to dense planting, making it a key trait involved in wheat plant architecture improvement. Only an appropriate tiller angle can ensure high crop yields. Genetic research on tiller angle in rice began as early as the 1990s, and a large number of tiller angle QTLs have been identified. However, compared to rice, genetic research on tiller angle in wheat is seriously lagging behind, and the number of tiller angle genes identified is very limited.

[0006] In view of this, in-depth exploration of genes related to plant height and tillering angle, and the use of genetic transformation technology to study and utilize functional genes can provide valuable genetic resources for the innovation of crop germplasm resources and genetic improvement, and will help promote the selection and breeding of new high-yield wheat varieties. Summary of the Invention

[0007] The present invention aims to provide a novel gene related to plant height and / or tillering angle, and its encoded protein and application. The technical problems to be solved are not limited to the technical subject described herein, and those skilled in the art can clearly understand other technical subjects not mentioned herein through the following description.

[0008] To achieve the above objectives, the present invention first provides a protein, which may be named TaRHT-A1p. The protein may be any of the following:

[0009] A1) a protein with the amino acid sequence shown in SEQ ID NO: 1;

[0010] A2) a protein having at least 80% identity with the protein of A1) and having the same function as the protein of A1) obtained by substitution, deletion, and / or addition of amino acid residues in the amino acid sequence of SEQ ID NO: 1;

[0011] A3) A fusion protein having the same function is obtained by connecting a tag to the N-terminus and / or C-terminus of A1) or A2).

[0012] The protein may be derived from wheat ( Triticum aestivum ).

[0013] The substitutions described in A2) may be conservative substitutions.

[0014] The connection in A3) can be achieved by direct peptide bond or through a linker.

[0015] To facilitate the isolation, purification, detection, and / or localization of the protein described in A1) or A2), a tag protein may be attached to its amino or carboxyl terminus. Such tags include, but are not limited to, GST (glutathione sulfhydryltransferase) tags, Trx (thioredoxin) tags, nitrogen utilization substrate A (NusA) tags, His-tags, MBP (maltose binding protein) tags, Flag tags, SUMO tags, HA (influenza hemagglutinin) tags, Myc tags, LacZ tags, CBD (cellulose binding domain) tags, bacteriophage T7 protein kinase (T7PK) tags, GFP (green fluorescent protein), CFP (cyan fluorescent protein), YFP (yellow-green fluorescent protein), mCherry (monomeric red fluorescent protein), or AviTag tags. The use of tags does not alter the function of the target protein, and those skilled in the art will know how to select an appropriate tag protein based on the desired purpose.

[0016] The present invention also provides a biomaterial, which may be any of the following:

[0017] B1) a nucleic acid molecule encoding the protein TaRHT-A1p;

[0018] B2) an expression cassette containing the nucleic acid molecule described in B1);

[0019] B3) a recombinant vector containing the nucleic acid molecule described in B1), or a recombinant vector containing the expression cassette described in B2);

[0020] B4) a recombinant microorganism containing the nucleic acid molecule described in B1), or a recombinant microorganism containing the expression cassette described in B2), or a recombinant microorganism containing the recombinant vector described in B3);

[0021] B5) A recombinant host cell containing the nucleic acid molecule described in B1), or a recombinant host cell containing the expression cassette described in B2), or a recombinant host cell containing the recombinant vector described in B3).

[0022] Furthermore, the biological materials can all express the nucleic acid molecules described in B1).

[0023] In the above biological material, the nucleic acid molecule in B1) may be a DNA molecule with a coding sequence or a nucleotide sequence such as that shown in SEQ ID NO: 2.

[0024] The nucleic acid molecule described herein can be DNA, such as cDNA, genomic DNA or recombinant DNA; the nucleic acid molecule can also be RNA, such as mRNA or hnRNA.

[0025] The nucleotide sequence shown in SEQ ID NO: 2 may be TaRHT-A1p The coding sequence (CDS) of the gene encodes the protein TaRHT-A1p with the amino acid sequence shown in SEQ ID NO: 1.

[0026] B1) The nucleic acid molecule may also include a nucleic acid molecule obtained by modifying the codon preference based on the nucleotide sequence shown in SEQ ID NO: 2.

[0027] Those skilled in the art can readily mutate the nucleotide sequence encoding the TaRHT-A1p protein using known methods, such as site-directed mutagenesis (including oligonucleotide primer-mediated site-directed mutagenesis, PCR-mediated site-directed mutagenesis, and cassette mutagenesis) or directed evolution (including error-prone PCR, DNA shuffling, and in vitro random priming recombination). Artificially modified nucleotide sequences that are 75% or more identical to the nucleotide sequence encoding the TaRHT-A1p protein (e.g., SEQ ID NO: 2) are derived from and equivalent to the nucleotide sequences of the present invention, as long as they encode the TaRHT-A1p protein and have the same function as the TaRHT-A1p protein.

[0028] In the above biological materials, the recombinant vector can be a cloning vector or an expression vector.

[0029] Furthermore, the recombinant vector can be a recombinant expression vector obtained by cloning the gene encoding the TaRHT-A1p protein into an expression vector (e.g., a prokaryotic expression vector, a eukaryotic expression vector, or a viral expression vector). Although the pWMB111 vector is used in the examples provided herein, the present invention is not limited to this specific vector. Those skilled in the art may utilize other suitable vectors, as long as the vector is capable of expressing the nucleic acid molecule encoding the TaRHT-A1p protein.

[0030] Existing plant expression vectors can be used to construct TaRHT-A1p Recombinant gene expression vectors. Such plant expression vectors include, but are not limited to, binary expression vectors (e.g., pBI series vectors (e.g., pBI121), pBIN series vectors (e.g., pBin19), pCAMBIA series vectors (e.g., pCAMBIA1300 vectors), pPZP series vectors, pGreen series vectors, pBIBAC series vectors, pSKI015 vectors, pSKI074 vectors, pRI101-AN vectors, etc.) and co-integration vectors (which can be constructed by inserting a segment homologous to a Ti plasmid or a segment thereof into an intermediate vector via homologous recombination or cloning). Such plant expression vectors contain elements required for exogenous gene expression, such as promoters, multiple cloning sites, terminators, and ribosome binding sites. Such plant expression vectors may also contain the 3' untranslated region of the exogenous gene, i.e., the polyadenylation signal and any other DNA segments involved in mRNA processing or gene expression. The polyadenylation signal can direct the addition of polyadenylic acid to the 3' end of the mRNA precursor. Examples of such signals include, but are not limited to, the transcribed untranslated regions of the 3' end of Agrobacterium crown gall-inducing (Ti) plasmid genes (such as the rouge synthase Nos gene) and plant genes (such as the soybean storage protein gene). When using Agrobacterium to introduce the gene encoding the protein TaRHT-A1p of the present invention, it is preferred to use an expression vector suitable for Agrobacterium, such as a binary vector or a modified vector thereof. Examples of such plant expression vectors include pBI121, pBIN19, pSMAB704, the pCAMBIA series of vectors, and the pGreen series of vectors.

[0031] use TaRHT-A1p When constructing a recombinant plant expression vector, any enhanced promoter or constitutive promoter can be added before the transcription start nucleotide, including but not limited to cauliflower mosaic virus (CaMV) 35S promoter, maize ubiquitin promoter (Ubi), rice actin promoter (Actin), Emu promoter, maize Adhl Gene promoter, rice rbcS Gene promoter, tomato rbcS Gene promoters and the potato pinⅡ gene promoter can be used alone or in combination with other plant promoters. Furthermore, when constructing plant expression vectors using the genes of the present invention, enhancers, including translational enhancers and transcriptional enhancers, can also be used. These enhancer regions can be the ATG start codon or an adjacent region start codon, but must be in frame with the coding sequence to ensure correct translation of the entire sequence. The translation control signals and start codons can be derived from a wide range of sources, both natural and synthetic. The translation initiation region can be derived from the transcriptional initiation region or a structural gene.

[0032] To facilitate the identification and screening of transgenic plant cells or plants, the plant expression vectors used can be modified to include, but are not limited to, genes encoding color-changing enzymes or luminescent compounds that can be expressed in plants (such as the GUS gene, luciferase gene, and GFP gene), antibiotic resistance genes (such as the kanamycin resistance gene kanr, the neomycin resistance gene neo, the hygromycin resistance gene hyg, the chloramphenicol resistance gene cat, the streptomycin resistance gene str, and the bleomycin resistance gene ble), or herbicide resistance genes (such as the bar gene, the glyphosate resistance marker gene epsps, and the chlorsulfuron resistance marker gene als). For the safety of transgenic plants, it is also possible to directly screen for transformed plants without adding any selectable marker genes.

[0033] In one or more embodiments of the present invention, the recombinant vector is pWMB111- TaRHT-A1p The recombinant vector pWMB111- TaRHT-A1p The recombinant expression vector is obtained by cloning a DNA fragment with the nucleotide sequence of SEQ ID NO: 2 into the BamHI recognition site of the pWMB111 vector while keeping other nucleotide sequences of the pWMB111 vector unchanged.

[0034] The present invention also provides the use of the protein TaRHT-A1p or the biomaterial in any of the following:

[0035] C1) Application in regulating plant strains;

[0036] C2) Application in regulating plant tillering angle;

[0037] C3) Application in regulating plant gravity perception or gravity response;

[0038] C4) Applications in breeding plants with altered plant height, tillering angle, gravity perception and / or gravity response;

[0039] C5) Applications in molecular breeding or germplasm improvement related to plant height, tillering angle, gravity perception and / or gravity response.

[0040] The application can be achieved by upregulating or downregulating the content and / or activity of the protein TaRHT-A1p.

[0041] Furthermore, the application may include upregulating the content and / or activity of the protein TaRHT-A1p (e.g., overexpression TaRHT-A1pThe application may also include down-regulating the content and / or activity of the protein TaRHT-A1p (e.g., knocking out or silencing the protein TaRHT-A1p). TaRHT-A1p genes) to increase plant height, reduce plant tillering angle, enhance plant gravity perception and / or enhance plant gravity response.

[0042] The present invention also provides a method for cultivating transgenic plants, which includes increasing the content and / or activity of the protein TaRHT-A1p in a target plant to obtain a transgenic plant, wherein the transgenic plant has a lower plant height than the target plant, a greater tillering angle than the target plant, and / or a weaker gravity perception ability or gravity response than the target plant.

[0043] In the above method, increasing the content and / or activity of the protein TaRHT-A1p in the target plant may be achieved by increasing the expression level of the gene encoding the protein TaRHT-A1p in the target plant.

[0044] Increasing the expression level of the gene encoding the protein TaRHT-A1p in the target plant can be achieved by at least one of the following methods:

[0045] (1) increasing the copy number of the gene encoding the protein TaRHT-A1p;

[0046] (2) expressing the gene encoding the protein TaRHT-A1p under the drive of a strong promoter;

[0047] (3) increasing the regulatory elements of the gene encoding the protein TaRHT-A1p to cause its overexpression, wherein the regulatory elements include enhancer elements, elements that improve mRNA stability, elements that enhance translation efficiency, and / or elements that enhance protein secretion;

[0048] (4) increasing the ribosome binding site of the gene encoding the protein TaRHT-A1p;

[0049] (5) codon optimization of the gene encoding the protein TaRHT-A1p;

[0050] (6) Upregulating the expression of the gene (the gene encoding the protein TaRHT-A1p) by changing epigenetic modifications such as DNA methylation or histone acetylation.

[0051] Furthermore, the method (2) can be achieved by replacing the natural promoter of the gene encoding the protein TaRHT-A1p with a strong promoter, or by operably linking a second promoter to the gene encoding the protein TaRHT-A1p.

[0052] The strong promoter includes, but is not limited to, T7 promoter, CaMV promoter, SV40 promoter, SFFV promoter, ubq promoter, ubi promoter, RBCS promoter, Actin promoter, Emu promoter, CYP450 promoter, Adhl promoter and pinⅡ promoter.

[0053] The enhancers described in method (3) include but are not limited to CMV enhancer, SV40 enhancer and RSV enhancer.

[0054] In the above method, increasing the expression level of the gene encoding the protein TaRHT-A1p in the target plant can be achieved by introducing the gene encoding the protein TaRHT-A1p into the target plant.

[0055] In the above method, the nucleotide sequence of the gene encoding the protein TaRHT-A1p may be as shown in SEQ ID NO: 2.

[0056] The present invention also provides a method for reducing the plant height of a target plant, increasing the tillering angle of a target plant, weakening the gravity perception ability of a target plant, or weakening the gravity response of a target plant, which method comprises increasing the content and / or activity of the protein TaRHT-A1p in the target plant.

[0057] The method for growing transgenic plants described herein may include the following steps:

[0058] (1) constructing a recombinant vector comprising a nucleic acid molecule encoding the protein TaRHT-A1p;

[0059] (2) introducing the recombinant vector constructed in step (1) into the target plant;

[0060] (3) Obtain transgenic plants through screening and identification.

[0061] Furthermore, the nucleic acid molecule encoding the protein TaRHT-A1p may be a DNA molecule whose coding sequence or nucleotide sequence is shown in SEQ ID NO: 2.

[0062] Furthermore, the above method may further comprise, after step (3), step (4): hybridizing the transgenic plant with the plant to be improved to obtain an offspring transgenic plant, wherein the offspring transgenic plant has substantially the same phenotype as the transgenic plant (i.e., the transgenic plant serving as the parent). The substantially identical phenotype may be a reduction in plant height, an increase in tillering angle, a weakened ability to sense gravity, and / or a weakened response to gravity compared to the plant to be improved.

[0063] Furthermore, the introduction method includes but is not limited to: Agrobacterium-mediated method, plant virus vector-mediated transformation method, gene gun method (also known as microparticle bombardment method or biological missile method), chemical stimulation method, electric shock method, liposome-mediated method, microinjection method, laser microbeam method, pollen tube channel method, ultrasonic method, air gun method and vortex method, etc.

[0064] Furthermore, the introduction method may be Agrobacterium-mediated method.

[0065] Furthermore, the Agrobacterium-mediated method may include the following steps: introducing the recombinant vector constructed in step (1) into Agrobacterium (such as Ca ion-induced transformation method, polyethylene glycol-mediated transformation method, metal cation-mediated transformation method, electroporation transformation method, phage transduction method, etc.) to obtain recombinant Agrobacterium, and infecting the callus tissue or explant of the target plant with the recombinant Agrobacterium; and inducing and culturing the positive callus tissue or explant obtained after identification to obtain regenerated plants.

[0066] The explants include but are not limited to seeds, roots, leaves, petioles, cotyledons, cotyledon petioles, hypocotyls, stem segments, stem apical meristems, epidermal parenchyma cells, tubers, stolon segments, embryonic suspension cells and protoplasts.

[0067] The screening and identification methods are known to those skilled in the art. For example, transformed transgenic plants (including transgenic offspring materials) can be identified by PCR detection, Southern hybridization, immunoblotting, Northern hybridization, enzyme-linked immunosorbent assay (ELISA), functional identification (testing the presence of selectable marker genes and target genes) and / or in situ hybridization.

[0068] Herein, the plant may be any of the following:

[0069] D1) Monocots or dicots;

[0070] D2) Grasses;

[0071] D3) Triticum genus;

[0072] D4) Wheat.

[0073] The regulation described herein can be upregulation (eg, increase or enhance) or downregulation (eg, decrease, reduce or attenuate).

[0074] Herein, the transgenic plants are understood to include not only first-generation transgenic plants obtained by introducing the gene encoding the protein TaRHT-A1p into target plants, but also their progeny. The transgenic plants include seeds, calli, whole plants and cells.

[0075] The inventors of the present invention have discovered the gene shown in SEQ ID NO: 2 after extensive and in-depth research. TaRHT-A1 A new allele of the gene, named TaRHT-A1p Gene. TaRHT-A1p Gene and wild-type gene ( TaRHT-A1 Compared with the gene), there is a difference in the 131st base of the CDS region (SEQ ID NO: 2), which is caused by TaRHT-A1 The G mutation in the gene is TaRHT-A1p The A in the gene, this base change causes the 44th glycine (G) in the corresponding amino acid sequence to mutate to glutamic acid (E). The mutation is located between the DELLA domain and the TVHYNP domain, adjacent to the DELLA domain. This mutation can regulate the plant height and tillering angle of wheat.

[0076] The present invention discovered and confirmed a new gene that regulates wheat plant height and tiller angle (wheat dwarf creeping allele) TaRHT-A1p ), and revealed for the first time TaRHT-A1p The application of the gene and its encoded protein TaRHT-A1p in regulating plant height, tillering angle, gravity perception and / or gravity response. The results of plant genetic transformation experiments and phenotypic analysis showed that by increasing the content and / or activity of the protein TaRHT-A1p in the target plant (e.g., overexpression), TaRHT-A1p Gene) can significantly reduce the plant height, increase the tillering angle of the plant, weaken the plant's gravity perception ability and / or weaken the plant's gravity response; accordingly, by reducing the content and / or activity of the protein TaRHT-A1p in the target plant (for example, knocking out or silencing TaRHT-A1p Genes) can significantly increase plant height, reduce plant tillering angle, enhance plant gravity perception and / or enhance plant gravity response.

[0077] The present invention has successfully created dwarf wheat with significantly reduced plant height and significantly increased tillering angle. Dwarf wheat is less likely to fall over, has stronger adaptability, can better withstand strong winds, reduce losses caused by natural disasters such as typhoons and tornadoes, and ensure harvest rate and quality. The increased tillering angle can improve the photosynthetic utilization efficiency of wheat, enhance field competitiveness, reduce pests and diseases, and increase yield. TaRHT-A1pGenes can be used to improve wheat plant types, thereby providing new ways to breed new high-yield wheat varieties and laying the foundation for increasing wheat yield and improving traits.

[0078] The present invention provides material accumulation and efficient and safe technical methods for wheat germplasm resource improvement and breeding, provides a new and good gene resource for wheat breeding, and is conducive to promoting the commercial breeding process of wheat.

[0079] Definition of terms

[0080] In the present invention, unless otherwise specified, the scientific and technical terms used herein have the meanings commonly understood by those skilled in the art. At the same time, in order to better understand the present invention, the definitions and explanations of relevant terms are provided below.

[0081] The term "expression cassette" generally refers to a nucleic acid construct containing nucleic acid elements sufficient to express a gene of interest. A typical expression cassette includes a promoter, a multiple cloning site (MCS), and a terminator. An expression cassette may also include the gene of interest, marker genes (such as TK, DHFR, CAT, and NEO), ribosome recognition and binding sites (SDs), transcription factor binding sites (TFBSs), enhancers, silencers, repressors, introns, poly(A) signal sequences, and / or mRNA splicing signal sequences. The various elements in an expression cassette can be linked directly or indirectly via linkers.

[0082] The term "vector" generally refers to a vehicle capable of transporting exogenous DNA or a gene of interest into host cells for amplification and / or expression. Such a vector can be either a cloning vector or an expression vector. A vector can be introduced into host cells via transformation, transduction, or transfection, enabling the amplification and / or expression of the genetic material it carries. Those skilled in the art can select an appropriate vector based on the objectives of the genetic engineering project and the properties of the recipient cells. The vectors include, but are not limited to, plasmids, phages (e.g., lambda phage or M13 phage), cosmids (i.e., cosmids), phagemids, shuttle vectors (e.g., yeast expression vectors), Ti plasmids, artificial chromosomes (e.g., yeast artificial chromosomes (YACs), bacterial artificial chromosomes (BACs), P1 artificial chromosomes (PACs), or Ti plasmid artificial chromosomes (TACs)), viral vectors (e.g., baculovirus vectors, retroviruses (including lentiviruses), adenoviruses, adeno-associated viruses, poxviruses, papillomaviruses, papillomaviruses (e.g., SV40), and herpesviruses (e.g., herpes simplex virus)). A vector may contain a variety of elements for controlling expression, including, but not limited to, promoter sequences, transcription initiation sequences, enhancer sequences, selection elements, and reporter genes. In addition, the vector may also contain a replication initiation site.

[0083] The term "microorganism" generally includes bacteria, viruses, fungi, actinomycetes, rickettsia, mycoplasma, chlamydia, spirochetes, algae, etc. For example, the bacteria may be from the genus Escherichia ( Escherichia sp. ) (such as Escherichia coli), Erwinia spp. ( Erwinia sp. ), Agrobacterium ( Agrobacterium sp. ) (such as Agrobacterium tumefaciens), Flavobacterium spp. ( Flavobacterium sp. ), Alcaligenes spp. ( Alcaligenes sp. ), Pseudomonas spp. ( Pseudomonas sp. ) and Bacillus spp. ( Bacillus sp. ) (such as Bacillus) and the like. The viruses may include rotavirus, baculovirus, retrovirus (such as lentivirus), adenovirus, adeno-associated virus, poxvirus, papillomavirus, influenza virus, papillomavirus (such as SV40) and herpes virus (such as herpes simplex virus) and the like. The fungus may be from the genus Saccharomyces ( Saccharomyces sp. ) (such as Saccharomyces cerevisiae, Methanol yeast, Pichia pastoris), Fusarium spp. ( Fusarium sp. ), Rhizoctonia spp. Rhizoctonia sp. ), Verticillium ( Verticillium sp. ), Penicillium ( Penicillium sp. ), Aspergillus ( Aspergillus sp. ) and Cephalosporium ( Cephalosporium sp. ) etc. The actinomycetes may be from the genus Streptomyces ( Streptomyces sp. ) (such as Streptomyces). The algae may be from Cyanophyta (such as cyanobacteria), Fucus ( Fucus sp. ), the genus A. Achnanthes sp. ), Cocoon algae ( Amphiprora sp. ), Diplocoriaceae ( Amphora sp. ), Fibrocystis spp. ( Ankistrodesmus sp. ), Astrophytum ( Asteromonas sp. ) and Chromophytes ( Boekelovia sp. )wait.

[0084] The term "host cell" is also called a recipient cell and generally refers to any type of cell that can be used to introduce a vector, such as plant cells and animal cells. The host cell can be understood to refer not only to a specific recipient cell, but also to the progeny of such a cell. Due to natural, accidental or intentional mutations and / or changes, the progeny may not be completely identical to the original parent cell, but are still included in the scope of host cells. Suitable host cells are known in the art, among which: the plant cell can be Arabidopsis thaliana ( Arabidopsis thaliana ),tobacco( Nicotiana tabacum ),corn( Zea mays )、Rice( Oryza sativa ),wheat( Triticum aestivum ) and other plant cells, but not limited thereto; the animal cells may be mammalian cells (such as Chinese hamster ovary cells (CHO cells), Chinese hamster ovary cell substrain (CHO-K1 cells), African green monkey kidney cells (Vero cells), SV40-transformed African green monkey kidney cells (COS cells), baby hamster kidney cells (BHK cells), mouse breast cancer cells (C127 cells), human embryonic kidney cells (HEK293 cells), human HeLa cells, fibroblasts, bone marrow cell lines, T cells or NK cells, etc.), avian cells (such as chicken or duck cells), amphibian cells (such as African clawed frogs ( Xenopus laevis ) cells or giant salamander ( Andrias davidianus ) cells), fish cells (such as grass carp, carp, rainbow trout or catfish cells), insect cells (such as Sf21 cells, Sf-9 cells or Hi-5 cells), etc., but are not limited thereto.

[0085] The term "recombinant vector" generally refers to a recombinant DNA molecule constructed by connecting an exogenous target gene to a vector in vitro. It can be constructed in any suitable manner, as long as the constructed recombinant vector can carry the exogenous target gene into the recipient cell and provide the exogenous target gene with the ability to replicate, integrate, amplify and / or express in the recipient cell.

[0086] The term "recombinant microorganism" generally refers to a microorganism whose genes have been manipulated and modified to produce a functionally altered recombinant microorganism. This can be achieved by introducing an exogenous gene of interest or a recombinant vector into the microorganism, or by directly editing the endogenous genes of the microorganism.

[0087] The term "recombinant host cell" generally refers to a recombinant host cell whose genes have been manipulated and modified to produce functionally altered recombinant host cells. This can include introducing an exogenous gene of interest or a recombinant vector into a host cell, or directly editing the endogenous genes of the host cell.

[0088] The term "linked" generally refers to the association of two or more molecules. The link can be covalent or non-covalent. As used herein, the link can be directly connected via a peptide bond or connected via a linker (joiner).

[0089] The term "identity" generally refers to the degree to which two (nucleotide or amino acid) sequences have identical residues at the same position in an alignment, and is typically expressed as a percentage. Identity, as used herein, may refer to amino acid sequence or nucleotide sequence identity. Two copies of identical sequences have 100% identity. Those skilled in the art will appreciate that amino acid sequence or nucleotide sequence identity can be determined using internet identity search sites, such as the BLAST page on the NCBI homepage. For example, amino acid sequence identity can be calculated using Advanced BLAST 2.1 by using blastp as the program, setting the Expect value to 10, all filters to OFF, using BLOSUM62 as the matrix, and setting the Gap existence cost, Perresidue gap cost, and Lambda ratio to 11, 1, and 0.85 (default values), respectively. The identity value (%) can then be obtained. Sequence analysis software (such as CLC Main Workbench and MegAlign™) can also be used for determination, for example, using the computer program BLAST with default parameters, particularly BLASTP or TBLASTN. The 75% or greater identity described herein may be 75%, 80%, 85%, 90% or greater identity. Herein, the 80% or greater identity may be at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or greater identity.

[0090] The term "conservative substitution" generally refers to the replacement of one amino acid residue with another amino acid residue having a side chain with similar physicochemical properties. For example, conservative substitutions can be made between amino acid residues with hydrophobic side chains (e.g., Met, Ala, Val, Leu, and Ile), between neutral hydrophilic side chain residues (e.g., Cys, Ser, Thr, Asn, and Gln), between acidic side chain residues (e.g., Asp, Glu), between basic side chain residues (e.g., His, Lys, and Arg), or between aromatic side chain residues (e.g., Trp, Tyr, and Phe). It is known in the art that conservative substitutions generally do not cause significant changes in protein conformational structure and do not substantially alter the biological activity of the protein. Conservative substitutions in protein sequences that are expected to have minimal or no effect on protein structure or function can be readily designed by those of ordinary skill in the art.

[0091] The term "overexpression" generally refers to increasing or upregulating the level and / or activity of a target protein or gene. Overexpression can be achieved through regulation at the gene level (e.g., gene replication, transcription, translation, post-transcriptional modification, and / or post-translational modification), or by promoting or increasing the content, activity, and / or function of the target protein at the protein level. The means of overexpression are not particularly limited, and those skilled in the art are familiar with numerous methods for achieving overexpression. For example, the nucleic acid molecule to be overexpressed or the nucleic acid molecule encoding the protein to be overexpressed can be placed under the control of a strong promoter; the copy number of one or more genes encoding the proteins described herein can also be increased; or the strength of ribosome binding sites or Kozak sequences can be increased, mRNA stability can be improved, codon usage can be altered, or inhibitory elements can be deleted.

[0092] The term "promoter" generally refers to a site specifically recognized and bound by RNA polymerase, located upstream of the transcription start site of a structural gene. It has strict directionality and initiates transcription. Because promoter strength determines transcription efficiency, different types of promoters can be used in genetic engineering to regulate the expression of key genes. Those skilled in the art are aware that using a strong constitutive promoter can overexpress a target gene. To further enhance target gene expression, multiple promoters can also be used in tandem.

[0093] The term "enhancer" generally refers to a DNA sequence that is located upstream or downstream of a structural gene and can also be located in an intron and can enhance gene transcription activity.

[0094] The term "regulatory element" generally refers to a DNA molecule with gene regulatory activity. Regulatory elements that function in plants include promoters, leader sequences, enhancers, introns, and 3'UTRs.

[0095] The term "operably linked" generally refers to the physical and / or functional connection of a DNA segment to another DNA segment in a manner that allows the segment to function in its intended manner. DNA encoding a gene product is operably linked to a regulatory element that can directly or indirectly regulate the transcription of the DNA. For example, when an enhancer is operably linked to DNA encoding a gene product, it can increase transcription of the DNA. The enhancer can be located upstream, downstream, or embedded within the coding region of the DNA.

[0096] The term "codon optimization" generally refers to a technique that increases protein expression levels in vivo by increasing the translation efficiency of target genes. Codon optimization typically involves redesigning genes to improve translation efficiency and, consequently, protein expression, by avoiding rare codons, utilizing preferred codons, simplifying mRNA secondary structure, optimizing repetitive sequences, eliminating restriction enzyme sites, and adjusting GC content.

[0097] The term "introduction" generally refers to the transfer of exogenous genes into recipient cells, such as eukaryotic recipient cells or prokaryotic recipient cells. The method of introduction is not particularly limited, and any known transformation method can be used as long as it can transfer the target gene (such as the DNA molecule of the present invention) into the recipient cell. The introduced DNA molecule can be a single copy or multiple copies. Introduction can be the integration of the exogenous gene into the host chromosome, or it can be expressed extrachromosomally by a plasmid. The introduction method may include any of the following: (1) introducing the target gene or a recombinant vector containing the target gene into the host bacteria by chemical transformation (such as Ca ion-induced transformation, polyethylene glycol-mediated transformation, or metal cation-mediated transformation, etc.) or physical transformation (such as electroporation transformation). (2) transducing the target gene into the host bacteria by phage transduction. (3) The target gene is transferred into the plant recipient cells by physical or chemical methods, such as gene gun method (also known as microparticle bombardment method or biological missile method), chemical stimulation method, electric shock method, liposome-mediated method, microinjection method, laser microbeam method, pollen tube channel method, ultrasound method, air gun method and vortex method, etc. (4) The target gene is transferred into the plant recipient cells by vector, such as Agrobacterium Ti plasmid vector (including Ti plasmid-derived vectors such as co-integration vector system and binary vector system) mediated method (Agrobacterium-mediated method), plant virus vector-mediated transformation method, etc.

[0098] The term "explant" generally refers to a portion of a plant used as in vitro culture material in plant tissue culture. After appropriate treatment and under suitable conditions, it can be regenerated into a whole plant. In practice, those skilled in the art will select the appropriate explant for transformation based on the specific plant.

[0099] The term "callus" generally refers to the newly formed tissue that forms on the surface of a wound after a local injury to the original plant. It is composed of living parenchyma cells and can originate from living cells in any tissue within the plant's organs. In plant tissue culture, it refers to a mass of unorganized, actively dividing parenchyma cells formed from an explant. Cultivating callus on an appropriate culture medium can induce it to form a whole plant. BRIEF DESCRIPTION OF THE DRAWINGS

[0100] Figure 1 for TaRHT-A1p Screening of positive seedlings of gene overexpressing plants.

[0101] Figure 2 for TaRHT-A1p Statistical analysis of the grain-filling phenotype, tillering angle, and plant height of gene-overexpressing plants.

[0102] Figure 3 for TaRHT-A1p Statistical analysis of jointing phenotype and tillering angle of gene overexpression plants.

[0103] Figure 4 Under normal light conditions TaRHT-A1p Results of gravity sensing experiments in gene-overexpressing plants at the seedling stage.

[0104] Figure 5 In dark conditions TaRHT-A1p Results of gravity sensing experiments in gene-overexpressing plants at the seedling stage.

[0105] Figure 6 Schematic diagram of the pWMB111 vector. DETAILED DESCRIPTION

[0106] The present invention will be further described in detail below in conjunction with specific embodiments. The examples provided are only for illustrating the present invention and are not intended to limit the scope of the present invention. The examples provided below can serve as a guide for further improvements by those skilled in the art and are not intended to limit the present invention in any way.

[0107] Unless otherwise specified, the experimental methods in the following examples are conventional methods and were performed according to the techniques or conditions described in the literature in the field or according to the product instructions. The materials and reagents used in the following examples, unless otherwise specified, were all commercially available.

[0108] The following examples were analyzed using GraphPad Prism statistical software. Results are presented as mean ± standard deviation using the Student's t test. P < 0.05 (*) indicates a statistically significant difference, P < 0.01 (**) indicates a statistically significant difference, and P < 0.001 (***) indicates a highly significant difference. Quantitative experiments in the following examples were performed in triplicate, unless otherwise noted, and the results are averaged.

[0109] The source of the dwarf, creeping mutant 2246 of the wheat Liangxing 99 variety used in the following examples is: Liangxing 99 is a wheat variety bred by Shandong Liangxing Seed Co., Ltd., with parental lines (Ji 91102 × Lumai 14 × PH85-16). 2246 is a dwarf mutant of Liangxing 99 obtained through EMS mutagenesis and was kindly provided by Professor Zhang Mingyi of the Shanxi Academy of Agricultural Sciences. This biological material is available to the public from the applicant. This material was used solely for repetition of experiments related to the present invention and is not for any other purpose.

[0110] The pWMB111 vector in the following examples was donated by Professor Ye Xingguo's research group at the Institute of Crop Science, Chinese Academy of Agricultural Sciences, and is described in the following literature: Riaz B, Chen HQ et al. Overexpression of Maize ZmC1 and ZmRTranscription Factors in Wheat Regulates Anthocyanin Biosynthesis in a Tissue-Specific Manner[J]. International Journal of Molecular Sciences, 2019, 20, 5806. See the vector map for details. Figure 6 .

[0111] Example 1 TaRHT-A1p Gene discovery and cloning

[0112] The inventors of this application conducted extensive and in-depth research. First, they used the dwarf creeping mutant 2246 of wheat Liangxing 99 as the research material and identified its phenotype. Then, they crossed it with the wild-type Liangxing 99 and Jing 411, constructed the segregating population of the F2 generation and its derived offspring, and conducted preliminary and fine mapping of the dwarf creeping gene. TaRHT-A1p The genes were listed as candidate genes. After screening and identification, genes that regulate wheat plant height and tiller angle were obtained. TaRHT-A1p The cloning and sequence of the gene are as follows:

[0113] Find wheat on the Ensembl Plants website TaRHT-A1 The sequence of the gene, according to the website TaRHT-A1 The base sequence of the gene was determined, and primers targeting its specific fragment were designed. Stems of mutant 2246 wheat were sampled at the heading stage, frozen in liquid nitrogen, and ground. RNA was extracted using the Trizol method, and reverse transcribed using the RNA as a template to obtain cDNA. PCR amplification was performed using the cDNA from the mutant 2246 wheat stem as a template, and the resulting amplified product was sequenced.

[0114] Sequencing results showed TaRHT-A1p The gene CDS is 1863 bp long and encodes 620 amino acid residues. The CDS nucleotide sequence is shown in SEQ ID NO:2, and its amino acid sequence is shown in SEQ ID NO:1. Compared with the wild type, TaRHT-A1p The 131st base mutation in the CDS region of the gene is caused by TaRHT-A1 The G mutation in TaRHT-A1p The A in the nucleotide sequence of the 44th base pair is changed from glycine (G) to glutamic acid (E). The mutation is located between the DELLA domain and the TVHYNP domain, adjacent to the DELLA domain.

[0115] Amino acid sequence of protein TaRHT-A1p:

[0116] MKREYQDAGGSGGGGGMGSSEDKMMVSAAAGEGEEVDELLAALEYKVRASDMADVAQKLEQLEMAMGMGGVGAGAAPDDSFATHLATDTVHYNPTDLSSWVESMLSELNAPPPPLPPAPQQLNASTSSTVTGGGYFDLPPSVDSSCSTYALRPIPSPAGAVGPADLSADSVRDPKRMRTGGSSTSSSSSSSSSLGGGARSSVVEAAPPVAAGANAPALPVVVVDTQEAGIRLVHALLACAEAVQQENFSAAEALVKQIPLLAASQGGAMRKVAAYFGEALARRVFRFRPQPDSSLLDAAFADLLHAHFYESCPYLKFAHFTANQAILEAFAGCRRVHVVDFGIKQGMQWPALLQALALRPGGPPSFRLTGVGPPQPDETDALQQVGWKLAQFAHTIRVDFQYRGLVAATLADLEPFMLQPEGEEDPNEEPEVIAVNSVFEMHRLLAQPGALEKVLGTVRAVRPRIVTVVEQEANHNSGTFLDRFTESLHYYSTMFDSLEGGSSGGPSEVSSGAAAAPAAAGTDQVMSEVYLGRQICNVVACEGAERTERHETLGQWRNRLGNAGFETVHLGSNAYKQASTLLALFAGGDGYKVEEKEGCLTLGWHTRPLIATSAWRLAAP (SEQ ID NO:1).

[0117] TaRHT-A1p Coding sequence (CDS) of the gene:

[0118]

[0119] Example 2 TaRHT-A1p Application of genes in regulating plant height and tillering angle in wheat

[0120] 1. Construction of recombinant expression vector

[0121] According to wheat TaRHT-A1p The CDS sequence of the gene was used to design primers using DNAman software. TaRHT-A1p The CDS sequence of the PCR product was recovered by gel recovery, and the obtained gel recovery product was connected with the pWMB111 vector cut with BamHI enzyme by seamless cloning enzyme, and transformed into Escherichia coli competent T1. Single colonies were picked for shaking and sent for testing. The bacteria with correct sequencing were selected for shaking and plasmid extraction. TaRHT-A1p Overexpression vector, named pWMB111- TaRHT-A1p . Recombinant vector pWMB111- TaRHT-A1p The recombinant expression vector is obtained by cloning a DNA fragment with the nucleotide sequence of SEQ ID NO: 2 into the BamHI recognition site of the pWMB111 vector while keeping other nucleotide sequences of the pWMB111 vector unchanged.

[0122] The designed primer sequences are as follows:

[0123] 111-A1p-F:5'-AGGTCGACTCTAGAGGATCCATGAAGCGCGAGTACCAGGA-3' (SEQ ID NO:3);

[0124] 111-A1p-R: 5'-AGCTCGGTACCCGGGGATCCTCACGGCGCGGCCAGGCGCC-3' (SEQ ID NO: 4).

[0125] 2. Obtaining genetically modified wheat

[0126] Immature wheat ears of Fielder wheat were washed three times in a clean bench with 5% sodium hypochlorite for 20 min each time, and then washed 4-5 times with sterile ddH2O. The intact embryos of the seeds were taken in the clean bench and placed in the corresponding liquid culture medium. The constructed wheat TaRHT-A1p overexpression vector (pWMB111- TaRHT-A1p ) was transformed into competent Agrobacterium (EHA105). Wild Fielder wheat embryos were used as experimental materials. The wheat genetic transformation was carried out using the Agrobacterium-mediated method. The specific steps are as follows:

[0127] Embryos were co-cultivated with Agrobacterium in WLS1 liquid medium (containing 1 / 10 Linsmaier and Skoog (LS) salts, 1 / 10 Murashige and Skoog (MS) vitamins, 10 g / L glucose, 0.5 g / L 2-(N-morpholino)ethanesulfonic acid (MES), and 100 μM acetosyringone (AS), pH 5.8) at room temperature for 5 min and then transferred to solid co-cultivation medium (WLS liquid medium supplemented with 0.85 mg / L silver nitrate, 1.25 mg / L copper sulfate pentahydrate, and 8 g / L agarose) with the scutellum facing up and incubated at 25°C in the dark for 2 days. After co-cultivation, the embryonic axis was removed with a scalpel, and the remaining scales were transferred to callus induction medium (containing LS salts, MS vitamins, 0.5 mg / L 2,4-D, 2.2 mg / L picloram, 0.85 mg / L silver nitrate, 100 mg / L ascorbic acid, 250 mg / L carbenicillin, 100 mg / L cefotaxime, 1.95 g / L MES, and 5 g / L agarose) and cultured under the same conditions for another 5 days. Subsequently, the tissue was transferred to selective medium (callus induction medium supplemented with 5 mg / L glufosinate (PPT, Sigma, no. 45520) and without cefotaxime) for callus induction. After 2 weeks, the calli were transferred to selective medium containing 10 mg / L PPT and cultured in the dark for 3 weeks to induce embryonic callus formation. Embryogenic calli were cultured on 1 / 2 MS medium (without zeatin) containing 5 mg / L PPT at 25°C under 100 μmol m⁻² s⁻¹ light conditions for differentiation. Regenerated shoots were transferred to rooting medium containing 5 mg / L PPT for elongation and rooting. After full root development, the plants were transplanted into pots and cultured in a growth chamber to obtain T0-generation transgenic seedlings.

[0128] 3. Molecular detection of genetically modified wheat

[0129] DNA was extracted from the transgenic lines obtained (CTAB method), and the TaRHT-A1p-OE transgenic wheat plants were qualitatively identified by PCR reaction. The electrophoresis test results showed that the target gene band could be amplified in the TaRHT-A1p-OE positive plants, while the wild Fielder control group had no target band (see Figure 1 , a sequence on the vector and a sequence on the target fragment were selected, with a total size of about 800 bp. This can indicate that the amplified band is the target gene on the introduced overexpression vector rather than the gene of the wheat itself). A total of 25 TaRHT-A1p-OE positive plants were identified ( TaRHT-A1pGene overexpression plants). Identification primers are as follows:

[0130] UBI-F (vector sequence): 5′-TAGCCCTGCCTTCATACGCT-3′ (SEQ ID NO: 5);

[0131] A1p-R (CDS sequence): 5′-TCCGCGGCAGAGAAGTTCTC-3′ (SEQ ID NO: 6).

[0132] Wheat genomic DNA was extracted using the CTAB method from fresh wheat seedling leaves. The main steps are as follows: First, a fresh wheat tissue sample was placed in a 2.0 mL centrifuge tube filled with a 6 mm diameter steel ball and quickly frozen in liquid nitrogen. The sample was then ground in a sample grinder until it reached a powdery consistency. Next, 800 μL of preheated CTAB extraction buffer was added to the ground sample, mixed thoroughly, and incubated in a 65°C water bath or oven for 30 minutes, gently shaking the mixture every 15 minutes. Following incubation, the sample was cooled to room temperature and 800 μL of a 24:1 chloroform / isoamyl alcohol mixture was added. The mixture was gently shaken for 5 minutes, allowed to stand at room temperature for 5 minutes, and then centrifuged at 12,000 rpm for 15 minutes at room temperature. After centrifugation, transfer 600 μL of the supernatant to a new 1.5 mL centrifuge tube. Add 0.8-1 volumes of isopropanol, gently invert to mix, and incubate at -20°C for at least 30 minutes (or overnight). Subsequently, centrifuge at 10,000 rpm at 4°C for 10 minutes and discard the supernatant. Add 1 mL of 75% ethanol to the pellet, gently shake to wash the DNA pellet, and centrifuge at 10,000 rpm at 4°C for 2 minutes. Repeat this wash step once, discard the supernatant, and air-dry the DNA pellet at room temperature. Finally, add an appropriate amount of ddH2O to dissolve the DNA and store at -20°C for subsequent analysis.

[0133] 4. Phenotypic identification of transgenic wheat

[0134] The tillering angle is the angle formed by the outermost tillers, and the plant height is the height from the base of the stem to the top of the tallest ear (excluding awn length).

[0135] 4-1. Tillering angle and plant height during the filling period

[0136] The constructed TaRHT-A1p-OE transgenic wheat lines 1# and 2# were planted in a greenhouse. The wild type Fielder was used as a control. The tiller angle and plant height phenotypes of the plants during the grain filling period were statistically analyzed. The results showed that the tiller angle of the TaRHT-A1p-OE plants was significantly greater than that of the wild type during the grain filling period (see Figure 2During the grain filling period, the average tillering angle of the wild-type plants was 22°, while that of the TaRHT-A1p-OE plants reached 45°, with the tillering angle of the 1# line being the most significant (see Figure 2 During the grain filling period, the average plant height of the wild type was 72 cm, while the average plant height of the TaRHT-A1p-OE plants was 48 cm. The plant heights of the two strains of TaRHT-A1p-OE plants were significantly lower than those of the wild type (see Figure 2 ). In summary, TaRHT-A1p The upregulation of gene expression levels resulted in an increase in wheat tillering angle and a decrease in plant height.

[0137] 4-2. Tillering angle during jointing stage

[0138] Since the phenotype of the constructed TaRHT-A1p-OE transgenic wheat line 1 was more obvious, we conducted a statistical analysis of its tiller angle phenotype at the jointing stage. The results showed that compared with the mature stage, the tiller angle of the TaRHT-A1p-OE plants increased more significantly at the jointing stage than that of the wild type, and appeared creeping (see Figure 3 At the jointing stage, the average tillering angle of wild-type plants was 20°, while that of TaRHT-A1p-OE plants reached 70°.

[0139] 4-3. Gravity perception or gravity reaction

[0140] The degree to which the aerial part perceives gravity is a key regulator of wheat tiller angle. Increased tiller angle in plants is often caused by defects in gravity response. To verify changes in the aerial part's gravity perception in TaRHT-A1p-OE seedlings, we conducted a gravity sensing experiment on stems during the seedling stage. The following method was used: Petri dishes were lined with seedling paper moistened with deionized water. Fielder seeds and seeds from two lines 1 and 2 of the constructed TaRHT-A1p-OE transgenic wheat were evenly distributed on the seedling paper. The seeds were incubated at room temperature for 1-2 days. Seeds of consistent growth were then sown at the top of a 15 ml centrifuge tube containing 0.4% agar. The tubes were placed vertically in a climatic chamber under a 16 h light / 8 h dark condition at 25°C for 3 days. The tubes were then placed horizontally for gravity stimulation. The tubes were incubated under 16 h light, 8 h dark, and complete darkness, with photographs taken every 12 h. The bending angle was measured by taking a picture of the wheat seedlings at each gravity-sensing treatment time point and then measuring the angle using ImageJ software.

[0141] The results showed that the bending angles of the stems of TaRHT-A1p-OE plants were significantly different from those of the wild type, both under light culture conditions and completely dark conditions, and were significantly smaller than those of the wild type plants (see Figure 4, Figure 5 ),therefore, TaRHT-A1p Increased gene expression can lead to a weakened perception of gravity in the stem, thereby reducing the bending angle of the stem and resulting in a creeping phenotype.

[0142] The present invention has been described in detail above. For those skilled in the art, without departing from the purpose and scope of the present invention, and without the need to carry out unnecessary experimental conditions, the present invention can be implemented in a wide range under equivalent parameters, concentrations and conditions. Although the present invention provides specific embodiments, it should be understood that further improvements can be made to the present invention. In short, according to the principles of the present invention, this application is intended to include any changes, uses or improvements to the present invention, including changes that depart from the disclosed scope in this application and are made using conventional techniques known in the art.

Claims

1. A protein, characterized in that The protein is any one of the following: A1) a protein with the amino acid sequence shown in SEQ ID NO: 1; A2) A fusion protein with the same function as A1) is obtained by connecting a tag to the N-terminus and / or C-terminus.

2. Biomaterial, characterized in that The biological material is any one of the following: B1) a nucleic acid molecule encoding the protein according to claim 1; B2) an expression cassette containing the nucleic acid molecule described in B1); B3) a recombinant vector containing the nucleic acid molecule described in B1), or a recombinant vector containing the expression cassette described in B2); B4) A recombinant microorganism containing the nucleic acid molecule described in B1), or a recombinant microorganism containing the expression cassette described in B2), or a recombinant microorganism containing the recombinant vector described in B3).

3. The biomaterial according to claim 2, characterized in that B1) The nucleic acid molecule is a DNA molecule whose coding sequence is shown in SEQ ID NO:

2.

4. Use of the protein according to claim 1, or the biomaterial according to claim 2 or 3, in any of the following: C1) Application in reducing plant strain height; C2) Application in increasing plant tillering angle; C3) Application in weakening plant gravity perception or gravity response; C4) Application in breeding plants with reduced plant height, increased tillering angle, reduced gravity perception and / or gravity response; The plant is wheat.

5. A method for cultivating transgenic wheat, characterized in that: The method comprises increasing the content of the protein according to claim 1 in wheat to obtain transgenic wheat, wherein the transgenic wheat has reduced plant height, increased tillering angle and / or weakened gravity perception or gravity response; The increasing of the content of the protein according to claim 1 in wheat is achieved by increasing the expression level of the gene encoding the protein in wheat.

6. The method according to claim 5, characterized in that The increasing of the expression level of the gene encoding the protein in wheat is achieved by introducing the gene encoding the protein according to claim 1 into wheat.

7. The method according to claim 6, characterized in that The nucleotide sequence of the gene encoding the protein is shown in SEQ ID NO:

2.

8. A method for reducing wheat plant height, increasing wheat tillering angle, weakening wheat's gravity perception ability, or weakening wheat's gravity response, characterized in that: The method comprises increasing the content of the protein according to claim 1 in wheat; the increasing the content of the protein according to claim 1 in wheat is achieved by increasing the expression level of the gene encoding the protein in wheat.

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