Temperature-mediated protein tafzp and application of encoding gene thereof

Abstract

The application discloses temperature-mediated protein TaFZP and application of a coding gene thereof, and belongs to the technical field of biotechnology.The amino acid sequence of the protein TaFZP provided by the application is shown as SEQ ID NO:2 or SEQ ID NO:4.It is proved by experiments that the temperature can change the ear type of the TaFZP mutant.The number of spikelets per ear can be significantly increased at 16 degrees in Fielder by knocking out TaFZP Therefore, the protein TaFZP has important application value in regulating the ear type of wheat and has a broad prospect in cultivating wheat varieties.

Description

Technical Field

[0001] This invention belongs to the field of biotechnology, specifically relating to the application of the temperature-mediated protein TaFZP and its encoding gene. Background Technology

[0002] With the increasing severity of global climate change, the cultivation of field crops is facing unprecedented challenges. Climate change has had a profound impact on crop yields and other aspects. How environmental temperature affects the growth and development of crop inflorescences is one of the hot topics in developmental biology research.

[0003] Wheat is one of the world's major food crops. Currently, with limited arable land, breeding varieties with longer ears and slightly more spikelets (e.g., 1-3 more spikelets per ear) to promote coordinated improvement of traits such as grain number and weight is an effective approach to high-yield wheat breeding. The wheat ear, the inflorescence organ, has always been an important trait of interest to wheat breeders. Wheat has a compound spike inflorescence, with several nodes on the rachis. Typically, each node bears a spikelet at its base, and each spikelet produces 3-10 florets. Changing the environmental temperature for wheat growth, thereby altering the number of spikelets at the base of each node (producing more spikelets or branching traits), could potentially increase the number of florets per ear, thus increasing the grain yield per plant. Summary of the Invention

[0004] The technical problem to be solved by this invention is to provide a new application for TaFZP protein in regulating plant spike type. The technical problem to be solved is not limited to the described technical subject matter; other technical subject matter not mentioned herein will be clearly understood by those skilled in the art through the following description.

[0005] To solve the above-mentioned technical problems, the present invention provides the following technical solutions: This invention provides applications of proteins or biological materials related to said proteins, said applications being any of the following: E1) Regulates plant spike-type traits; E2) Preparation of products that regulate plant spikelet traits; E3) Plant breeding; The biological material is a substance that regulates the expression of the gene encoding the protein or a substance that regulates the content of the protein. The protein is any one of the following: B1) The amino acid sequence is that of the protein shown in SEQ ID NO:2 and / or SEQ ID NO:4; B2) A protein obtained by substituting and / or deleting and / or adding amino acid residues of the protein described in B1) that has more than 80% identity with and the same function as the protein shown in B1). B3) A fusion protein obtained by attaching a protein tag to the N-terminus and / or C-terminus of B1) or B2).

[0006] In the above applications, the protein tag refers to a polypeptide or protein fused with a target protein using in vitro DNA recombination technology for expression, detection, tracing, and / or purification of the target protein. The protein tag may be a Poly-Arg tag, Strep-tag II tag, Flag tag, His tag, MBP tag, HA tag, myc tag, GST tag, and / or SUMO tag, etc.

[0007] In the above applications, identity refers to the identity of amino acid sequences. The identity of amino acid sequences can be determined using homology search sites on the Internet, such as the BLAST page on the NCBI homepage. For example, in Advanced BLAST 2.1, using blastp as the program, setting the Expect value to 10, setting all filters to OFF, using BLOSUM62 as the matrix, setting the Gap existence cost, Per residue gap cost, and Lambda ratio to 11, 1, and 0.85 (default values) respectively, and performing an identity search on a pair of amino acid sequences to calculate the identity value (%), then the identity value can be obtained.

[0008] In the above applications, the 80% or more of identity can be at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 95%, 96%, 98%, 99%, or 100% identity.

[0009] In the above applications, the protein is derived from wheat.

[0010] In the above applications, the regulation of plant spike type traits is any one of the following: e1) Number of spikelets in the plant; e2) Number of plant florets.

[0011] In the above applications, the biomaterial is any one of the following: C1) Nucleic acid molecules that inhibit, reduce, or downregulate the expression of the genes encoding the aforementioned proteins; C2) expresses the gene encoding the nucleic acid molecule described in C1); C3) contains an expression cassette containing the gene described in C2); C4) A recombinant vector containing the gene described in C2), or a recombinant vector containing the expression cassette described in C3); C5) Recombinant microorganisms containing the gene described in C2), or recombinant microorganisms containing the expression cassette described in C3), or recombinant microorganisms containing the recombinant vector described in C4); C6) A transgenic plant cell line containing the gene described in C2), or a transgenic plant cell line containing the expression cassette described in C3), or a transgenic plant cell line containing the recombinant vector described in C4); C7) Transgenic plant tissue containing the gene described in C2), or transgenic plant tissue containing the expression cassette described in C3), or transgenic plant tissue containing the recombinant vector described in C4); C8) A transgenic plant organ containing the gene described in C2), or a transgenic plant organ containing the expression cassette described in C3), or a transgenic plant organ containing the recombinant vector described in C4).

[0012] In the aforementioned biological materials, the expression cassette containing nucleic acid molecules described in B3) refers to DNA capable of expressing the RNA molecules described above in host cells. The expression cassette containing nucleic acid molecules described in B9) refers to DNA capable of expressing the proteins described above in host cells. The expression cassette may also include single-stranded or double-stranded nucleic acid molecules containing all the regulatory sequences necessary for expressing the DNA of any of the aforementioned proteins or RNA molecules. The regulatory sequences, under compatible conditions, can guide the coding sequence to express the DNA of any of the aforementioned proteins or RNA molecules in suitable host cells. The regulatory sequences include, but are not limited to, leader sequences, polyadenylated sequences, propeptide sequences, promoters, signal sequences, and transcription terminators. At a minimum, the regulatory sequences must include a promoter and termination signals for transcription and translation. To introduce specific restriction enzyme sites of the vector for linking the regulatory sequences to the coding region of the nucleic acid sequence encoding the protein or the DNA of the RNA molecule, a regulator-linked regulatory sequence may be provided. The regulatory sequence may be a suitable promoter sequence, i.e., a nucleic acid sequence that can be recognized by the host cell expressing the nucleic acid sequence. The promoter sequence contains a transcriptional regulatory sequence mediating the DNA expression of the protein or the RNA molecule. The promoter can be any nucleic acid sequence that is transcriptionally active in the selected host cell, including mutated, truncated, and heterozygous promoters, and can be derived from genes encoding extracellular or intracellular proteins that are homologous or heterologous to those of the host cell. The regulatory sequence can also be a suitable transcription termination sequence, i.e., a sequence that can be recognized by the host cell and thus terminate transcription. The termination sequence is operatively linked to the 3' end of the nucleic acid sequence encoding the protein or the DNA of the RNA molecule. Any terminator that can function in the selected host cell can be used in this invention. The regulatory sequence can also be a suitable leader sequence, i.e., an untranslated region of mRNA that is crucial for translation in the host cell. The leader sequence is operatively linked to the 5' end of the nucleic acid sequence encoding the protein or the DNA of the RNA molecule. Any leader sequence that can function in the selected host cell can be used in this invention. The regulatory sequence can also be a signal peptide coding region, which encodes an amino acid sequence linked to the amino terminus of a protein, capable of guiding the DNA encoding the protein or the RNA molecule into the cellular secretion pathway. Signal peptide coding regions that can guide the expressed protein or the DNA of the RNA molecule into the secretion pathway of the host cell used can be used in this invention. Adding regulatory sequences that can modulate the expression of proteins or RNA molecules according to the growth status of the host cell may also be necessary. Examples of regulatory sequences are systems that respond to chemical or physical stimuli (including in the presence of regulatory compounds), thereby turning gene expression on or off. Other examples of regulatory sequences are those that enable gene amplification.

[0013] In the aforementioned biological materials, the carrier may be a plasmid, a granule, a bacteriophage, or a viral vector.

[0014] In the above-mentioned biological materials, the microorganisms may be yeast, bacteria, algae or fungi, such as Agrobacterium.

[0015] Among the aforementioned biological materials, the transgenic plant cell lines do not include propagation materials.

[0016] In the above applications, the nucleic acid molecule described in B1) is a gRNA that targets the aforementioned protein-coding gene.

[0017] The present invention also provides a method for regulating spike type traits of plants, comprising the following steps: the method includes introducing a substance into a target plant that inhibits, reduces, or downregulates the expression of the gene encoding the aforementioned protein or inhibits, reduces, or downregulates the activity and / or content of the aforementioned protein, thereby altering the spike type trait of the target plant, wherein the target plant contains the gene encoding the aforementioned protein.

[0018] The present invention also provides a method for cultivating plants with altered spikelet type traits, comprising the following steps: the method includes introducing a substance that inhibits, reduces, or downregulates the expression of the gene encoding the aforementioned protein or inhibits, reduces, or downregulates the activity and / or content of the aforementioned protein into a target plant to obtain a plant with altered spikelet type traits, wherein the target plant contains the gene encoding the aforementioned protein.

[0019] Gene knockout refers to the phenomenon of inactivating a specific target gene through homologous recombination. Gene knockout inactivates a specific target gene by altering its DNA sequence.

[0020] Gene silencing refers to the phenomenon of preventing or reducing gene expression without damaging the original DNA. Gene silencing presupposes no change in the DNA sequence, resulting in the absence or reduction of gene expression. Gene silencing can occur at two levels: transcriptional silencing due to DNA methylation, heterochromatinization, and position effects; and post-transcriptional gene silencing, which inactivates the gene at the post-transcriptional level through specific inhibition of target RNA. This includes antisense RNA, co-suppression, gene quelling, RNA interference (RNAi), and microRNA (miRNA)-mediated translational repression.

[0021] In the above method, the introduction of substances into the target plant that inhibit, reduce, or downregulate the expression of the gene encoding the aforementioned protein or inhibit, reduce, or downregulate the activity and / or content of the protein includes introducing a gene knockout vector into the target plant targeting nucleotides 381 to 402 of SEQ ID NO:1 or nucleotides 390 to 411 of SEQ ID NO:3, or targeting nucleotides 512 to 533 of SEQ ID NO:1 or nucleotides 521 to 542 of SEQ ID NO:3.

[0022] In the above method, the regulation of plant spike type traits can be any one of the following: e1) Number of spikelets in the plant; e2) Number of plant florets.

[0023] In the above applications or methods, the plant can be a plant that grows at room temperature or a plant subjected to low-temperature stress. The low temperature can be 16°C.

[0024] In the above applications or methods, the plant is any one of the following: G1) Dicotyledons or monocotyledons; G2) Plants of the order Poales; G3) Gramineae plants; G4) Plants of the Triticum genus; G5) Wheat.

[0025] Experiments of this invention demonstrate that protein genes can be simultaneously edited in wild-type wheat Fielder. TaFZP-A and TaFZP-D At 25 degrees Celsius, the mutant exhibited a multi-spike phenotype; at 16 degrees Celsius, the mutant exhibited a spikelet branching phenotype, and at 16 degrees Celsius... TaFZ The P mutant showed a significantly increased number of spikelets per spike. Therefore, the protein TaFZP has important application value in temperature-mediated regulation of wheat spike type traits and has broad prospects in wheat variety breeding. Attached Figure Description

[0026] Figure 1 for TaFZP Genotypic analysis of gene knockout mutants. "-" indicates a missing nucleotide, and the number indicates the number of missing nucleotides. Underlined sites indicate NGG sites.

[0027] Figure 2 Analysis of spike type in wheat material Zang734 at different temperatures.

[0028] Figure 3 For different temperatures TaFZPAnalysis of spikelet type and agronomic traits in gene knockout mutants; Bar = 3cm. A: Phenotypic analysis of Fielder and its gene-edited mutants at 25°C. B: Phenotypic analysis of Fielder and its gene-edited mutants at 16°C. C: Comparative analysis of the number of spikelets per spike in Fielder and its gene-edited mutants at 25°C and 16°C. D: Comparative analysis of the number of florets per spike in Fielder and its gene-edited mutants at 25°C and 16°C. E: Comparative analysis of ectopic spikelets (ES) and ectopic inflorescences (EI) per spike in gene-edited mutants at 25°C and 16°C. ES: ectopic spikelet; EI: ectopic inflorescence. Detailed Implementation

[0029] The present invention will now be described in further detail with reference to specific embodiments. The given embodiments are merely illustrative of the invention and not intended to limit its scope. The embodiments provided below can serve as a guide for further improvements by those skilled in the art and do not constitute a limitation on the invention in any way.

[0030] Unless otherwise specified, the experimental methods used in the following examples are conventional methods, performed according to the techniques or conditions described in the literature in this field or according to the product instructions. Unless otherwise specified, the materials and reagents used in the following examples are commercially available.

[0031] In the quantitative experiments in the following examples, three replicate experiments were set up, and the average value of the results was taken.

[0032] The pMETaU6.1 and pLGY-E003 plasmids are described in the non-patent literature “Yongxing Chen, Huixin Xiao, Yuange Wang, Wenling Li, Lingchuan Li, Lingli Dong, Xuebo Zhao, Miaomiao Li, Ping Lu, Huaizhi Zhang, Guanghao Guo, Keyu Zhu b, Beibei Li, Lei Dong, Peng Chen, Shuming Wu c, Yunbo Jiang, Fei Lu, Chengguo Yuan, Zhiyong Liu, Yusheng Zhao, Qiuhong Wu, WPA1 encodes a vWA domain protein that regulates sweat plant architecture. 2024:12 992-1000. https: / / doi.org / 10.1016 / j.cj.2024.05.008”. They are publicly available from the Institute of Genetics and Developmental Biology, Chinese Academy of Sciences. This biological material is only for repeating experiments related to this invention and may not be used for other purposes.

[0033] Wild-type wheat Fielder: described in the non-patent literature “Yuange Wang, Fei Du, Jian Wang, Ke Wang, Caihuan Tian, ​​Xiaoquan Qi, Fei Lu, Xigang Liu, Xingguo Ye, Yuling Jiao. Improving bread wheat yield through modulating an unselected AP2 / ERF gene. Nat Plants. 2022 Aug;8(8):930-939. doi: 10.1038 / s41477-022-01197-9. Epub 2022 Jul 18.”, is available to the public from the Institute of Genetics and Developmental Biology, Chinese Academy of Sciences. This biological material is only for repeating the relevant experiments of this invention and cannot be used for other purposes.

[0034] Wheat material Zang734: described in the non-patent literature “FRIZZY PANICLE defines a regulatory hub for simultaneously controlling spikelet formation and awn elongation in breadwheat”, is available to the public from the Institute of Genetics and Developmental Biology, Chinese Academy of Sciences. This biomaterial is only for repeating the relevant experiments of this invention and may not be used for other purposes.

[0035] The following examples used GraphPad Prism 8 statistical software to process the data. The experimental results are expressed as mean ± standard deviation. One-way ANOVA was used, and P < 0.05 (*) indicates that there is a significant difference.

[0036] Example 1, T2 generation TaFZP Phenotypic identification of gene knockout wheat under low temperature stress I. Effects of different temperatures on ear type of wheat variety Zang734 Zang734 is a Tibetan local species that exhibits the trait of triple spikelets. Map-based cloning revealed that this trait is caused by... WFZP-A and WFZP-D Caused by mutation.

[0037] The experiment was repeated three times, with each repetition as follows: The wheat material Zang734 was planted at 25°C and 16°C (350 μmol photons m) respectively. -2 s -1 (At a humidity of 60-70%), the wheat ears were photographed, and the number of spikelets per ear and the branching index were counted. Branching index = total number of spikelets per ear / number of nodes on the rachis per ear. The results are as follows Figure 2 As shown, low temperature can significantly increase the number of spikelets per ear in wheat material Zang734 ( Figure 2 In the middle F, Spikelet number per spike) and branching index ( Figure 2 (G, Branch index).

[0038] II. Effects of different temperatures on wheat materials tafzp-ad#1 and tafzp-ad#2 Influence of ear type tafzp-ad#1 and tafzp-ad#2 are derived from Example 2.

[0039] The experiment was repeated three times, with each repetition as follows: 1) tafzp-ad#1+16℃ group: The mutant wheat seeds tafzp-ad#1 were germinated at 20 degrees Celsius. After germination, the seeds were transplanted into flowerpots and planted in a plant artificial climate chamber at 16℃ with long day and light intensity PPFD=350umolm. -2 s -1 Each treatment involves 15-20 seedlings. Photographs are taken 20 days after flowering to record the spikelet number per spikelet, and the number of ES and EI spikelets per spikelet. The procedure for counting ES and EI spikelets per spikelet is as follows: Each rachis segment bears one main spikelet. If there are additional spikelets on one or both sides, these can be defined as ES. If there are inflorescence branches, they can be defined as EI. (e.g.) Figure 3 (As shown in B).

[0040] 2) tafzp-ad#2+16℃ group: The difference between this group and the tafzp-ad#1+16℃ group is that the tafzp-ad#2 seed is used instead of the tafzp-ad#1 seed. The rest of the operation is the same as the tafzp-ad#1+16℃ group.

[0041] 3) Wild-type control Fielder+16℃ group: The difference between this group and the tafzp-ad#1+16℃ group is that the wild-type control Fielder seeds are used instead of the tafzp-ad#1 seeds. The other operations are the same as the tafzp-ad#1+16℃ group.

[0042] 4) tafzp-ad#1+25℃ group: The difference between this group and the tafzp-ad#1+16℃ group is that 25℃ is replaced by 16℃, and the rest of the operation is the same as the tafzp-ad#1+16℃ group.

[0043] 5) tafzp-ad#1+25℃ group: The difference between this group and the tafzp-ad#1+25℃ group is that the tafzp-ad#2 seed is used instead of the tafzp-ad#1 seed. The rest of the operation is the same as the tafzp-ad#1+25℃ group.

[0044] 6) Wild-type control Fielder+25℃ group: The difference between this group and the tafzp-ad#1+25℃ group is that the wild-type control Fielder seeds are used instead of the tafzp-ad#1 seeds. The other operations are the same as the tafzp-ad#1+25℃ group.

[0045] The results are as follows Figure 3 As shown, compared with the control Fielder, the knockout mutants tafzp-ad#1 and tafzp-ad#2 significantly increased the number of spikelets per spike. P<0.001), and the number of ES and EI per ear increased significantly ( P <0.001), the increase in spikelet number at 16 degrees is mainly due to the production of additional spikelets and inflorescences.

[0046] The above results indicate that the temperature-mediated protein TaFZP has important application value in regulating wheat spike traits (such as spikelet number and floret number).

[0047] Example 2: Knockout of wheat TaFZP Genes influence wheat's response to low temperature stress 1. Wheat TaFZP Genetic information wheat A genome TaFZP The gene sequence is SEQ ID NO:1, and the amino acid sequence of its encoded protein is SEQ ID NO:2; in the wheat D genome TaFZP The gene sequence is SEQ ID NO:3, and the amino acid sequence of the encoded protein is SEQ ID NO:4.

[0048] 2. Primer design and amplification For target genes TaFZP-A and its homologous genes TaFZP-D Gene editing targets were designed based on conserved sequences, and the targets are as follows: T1: 5'-CCTCGCGCCGTTCCACGCGCAG-3' (corresponding to nucleotides 381 to 402 of SEQ ID NO:1 and nucleotides 390 to 411 of SEQ ID NO:3). T2: 5'-GCTACCACCAGCAGGGCCCGGG-3' (corresponding to nucleotides 512 to 533 of SEQ ID NO:1 and nucleotides 521 to 542 of SEQ ID NO:3); Forward primer F and reverse primer R were designed and synthesized. PCR amplification was performed using the intermediate vector pMETaU6.1 as a template. The PCR amplification system is shown in Table 1, and the PCR amplification program is shown in Table 2. The obtained PCR product contained two target sequences and the TaU6.1 promoter sequence.

[0049] The primer sequences are as follows: F: 5'-ccgaggtctcgggcgCCTCGCGCCGTTCCACGCGCAGgtttcagagctatgctggaaac -3'; R:5'- acctcggtctccaaacCCCGGGCCCTGCTGGTGGTAGCcaagtctgatgcagcaagc -3'.

[0050] Table 1. PCR amplification system

[0051] Table 2. PCR Amplification Procedure

[0052] 3. Ligation of PCR products with gene editing vectors The gene editing backbone vector pLGY-E003 was digested with Bsa1, and the vector was recovered from the gel. The PCR product was ligated with the aforementioned digested vector to obtain the recombinant plasmid pLGY-E003-TaFZP. - sgRNA. The ligation reaction conditions were as follows: 37℃ for 5 min, 16℃ for 10 min, for a total of 60 cycles; then 16℃ for 1 h. The ligation reaction system is shown in Table 3. Recombinant vector pLGY-E003-TaFZP - sgRNA can produce Cas9 and sgRNA targeting T1 and T2.

[0053] The nucleotide sequence of the sgRNA targeting T1 is as follows: 5'-CCUCGCGCCGUUCCACGCGCAG-3'.

[0054] The nucleotide sequence of the sgRNA targeting T2 is as follows: 5'-GCUACCACCAGCAGGGCCCGGG-3'.

[0055] Table 3. Connection Reaction System

[0056] 4. Agrobacterium-mediated transformation The recombinant plasmid pLGY-E003-TaFZP - sgRNA was transformed into Agrobacterium tumefaciens strain EHA105 to obtain the recombinant plasmid pLGY-E003-TaFZP. - Recombinant Agrobacterium tumefaciens EHA105 / pLGY-E003-TaFZP containing sgRNA - sgRNA.

[0057] Recombinant Agrobacterium EHA105 / pLGY-E003-TaFZP - Wheat Fielder (recipient) was infected with sgRNA, and T0 transgenic wheat was obtained by following the procedure: Agrobacterium-mediated genetic transformation was performed using recombinant strain EHA105 / pLGYE-3-Tavrs3. - sgRNA was transferred into the fielder embryo shield of wild-type wheat and then cultured.

[0058] 5. Identification of positive plants The identification methods are as follows: Ten T0 generation plants were randomly selected. TaFZP The specific steps for PCR testing of the proposed genetically modified wheat are as follows: 1) Total DNA was extracted from the leaves of 10 T0 generation transgenic wheat seedlings using the CTAB genomic DNA extraction method. The DNA content in the leaves of each T0 generation transgenic wheat was approximately 200 ng / μL.

[0059] 2) Use PCR technology to obtain fragment sequences, including gene editing target sites, from each T0 generation of transgenic wheat.

[0060] The PCR primers were: forward primer: 5'-CTAGGCTGGAGCAGTAGTATAGTAG-3' and reverse primer: 5'-TCTCCAAACAAACACTACTACCTAG-3'.

[0061] The reaction system is 20 μL, consisting of 10 μL of SYBR. ® PremixExTaq TM 0.5 μL of 10 μM forward primer, 0.5 μL of 10 μM reverse primer, and 1 μL of T0 metabolite. TaFZP It consists of genomic DNA from genetically modified wheat and 8.0 μL of nuclease-free water.

[0062] Reaction program: 94℃ pre-denaturation for 3 min; 95℃ denaturation for 3 sec; 55℃ annealing for 30 sec; 35 cycles.

[0063] 3) Sanger sequencing to detect mutants.

[0064] The PCR products were subjected to Sanger sequencing to detect gene editing. The mutants used in the experiment in Example 1 were named tafzp-ad#1 and tafzp-ad#2.

[0065] Compared to wild-type Fielde, the A and B genomes of tafzp-ad#1 and tafzp-ad#2 TaFZP See gene sequence changes Figure 1 As shown.

[0066] SEQ ID NO:1 5'-ATGAGCTCTCGCAGCAGCAGCGGCGGCGGCGGGTGCTCCCCAGATGA TGGCCTTCTCGGAGCATTCGCTGCCGAAGCCGATCGCCGGTCACCCGCAGCCGCAGCCGTCCCCGCCGTCGTCGCCGAGCGAGCGGCCGGCGGCGCGCGGCAGGCGGCGCGCGCAGGAGCCCGGGCGCTTCCTGGGCGTGCGCCGGCGGCCGTGGGGCCGGTACGCGGCCGAGATACGCGACCCGACCACCAAGGAGCGGCACTGGCTCGGCACCTTCGACACGGCGCAGGAGGCCGCCCTGGCCTACGACCGCGCCGCGCTCTCCATGAAGGGCGCGCAGGCGCGCACCAACTTCGTCTACGCGCACGCCGCCTACAACAACTACCCGCCCTTCCTCGCGCCGTTCCACGCGCAGCACCAGCCCGCCGCCTACGCCGCGTCCTCGGCCATGCCGTACGGCGGCCAGCAGCAGCACGCGGGCGCGGGGCCGCCGCACATTGGCAGCTCGTACCACCACGGCCACGGCTACCACCAGCAGGGCCCGGGCGAGTGTTCCATGCCGGTGCCCAGTGCCGCGGATCACGGCGCCAGCGGCCCGATGGACGTGCGCGGCAGCAGCGGCCACGACTTCCTCTTCCCCAGCGCCGACGACAACTCCGGGTACCTGAGCAGCGTGGTGCCGGAGAGCTGCCTCCGGCCCCGCGGCGGCGACCTGCAGGACGCGCGGCGCTACTCCGTGTCCGACGCCGACGCCTACGGGCTGGGCCTCCGGGAGGACGTGGACGACCTGGCGACGATGGTGGCCGGCTTCTGGGGCGGCGCCGACGCGCCGTACGGCGGCGGCCACGACATGGTCGCCTCGTCGCAGGGCTCGGACAACGGCTACTCCCCCTTCAGCTTCCTCTCCCACTGA-3’。

[0067] SEQ ID NO:2 MSSRSSSGGGGASQMMAFSEHSLPKPIAGHPQPQPSPPSSPSERPAARGRRRAQEPGRFLGVRRRPWGRYAAEIRDPTTKERHWLGTFDTAQEAALAYDRAALSMKGAQARTNFVYAHAAYNNYPPFLAPFHAQHQPAAYAASSAMPYGGQQQHAGAGPPHIGSSYHHGHGYHQQGPGECSMPVPSAADHGASGPMDVRGSSGHDFLFPSADDNSGYLSSVVPESCLRPRGGDLQDARRYSVSDADAYGLGLREDVDDLATMVAGFWGGADAPYGGGHDMVASSQGSDNGYSPFSFLSH。

[0068] SEQ ID NO:3: 5’-ATGAGCATCCGCAGCAGCAGCGGCGGCAGCGGCGGCGGCCATGCCTCCCAGATGATGGCGTTCTCGGAGCATTCGCTGCCGAAGCCGATCGCCGGCCACCCGCAGCCGCAGCCGTCCCCGCCGTCGTCTCCGAGCGAGAGGCCGGCGCCGCGTGGCAGGCGGCGCGCGCAGGAGCCCGGGCGCTTCCTGGGCGTGCGCCGGCGGCCGTGGGGCCGGTACGCGGCGGAGATACGCGACCCGACCACCAAGGAGCGGCACTGGCTCGGCACCTTCGACACGGCGCAGGAGGCCGCCCTGGCCTACGACCGCGCCGCGCTCTCCATGAAGGGCGCGCAGGCGCGCACCAACTTCGTCTACGCGCACGCCGCCTACAACAACTACCCGCCCTTCCTCGCGCCGTTCCACGCGCAGCAGCAGCCCGCCGCCTACGCGTCCTCGACCATGCCGTACGCCGGCCAGCAGCACGCGGCGCCGCACATTGGCAGCTCGTACCACCACGGCCACGGCCACGGCGGCCTCGGCTACCACCAGCAGGGCCCGGGCGCCGGCGCGGGCGAGTGCTCCATGCCGGTGCCCAATGCCGCCGATCACGGCGCCAGCAGCCCGATGGACGTGCGCGGCAGCAGCGGCCACGACTTCCTCTTCCCCAGCGCCGACGACAACTCCGGGTACCTGAGCAGCGTGGTGCCGGAGAGCTGCCTCCGGCCCCGCGGCGGCGACCTGCAGGACGCGCGGCGCTACTCCGTGTCCGACGCCGACGCCTACGGGCTGGGCCTCCGGGAGGACGTGGACGACCTGGCGTCCATGGTGGCCGGCTTCTGGGGCGGCGCCGACGCGGCGTACGGCGGGTTCGCCCCCGCGAACGGCGGCGGCCACGACATGGTCGCCTCGTCGCAGGGCTCCGACAACGGCTACTCCCCCTTCAGCTTCCTCTCCCACTGA-3’。

[0069] SEQ ID NO:4: MSIRSSSGGSGGGHASQMMAFSEHSLPKPIAGHPQPQPSPPSSPSERPAPRGRRRAQEPGRFLGVRRRPWGRYAAAEIRDPTTKERHWLGTFDTAQEAALAYDRAALSMKGAQARTNFVYAHAAYNNYPPFLAPFHAQQQPAAYASSTMPYAGQQHAA PHIGSSYHHGHGHGGLGYHQQGPGAGAGECSMPVPNAADHGASSPMDVRGSSGHDFLFPSADDNSGYLSSVVPESCLRPRGDLQDARRYSVSDADAYGLGLREDVDDLASMVAGFWGGADAAYGGFAPANGGGHDMVASSQGSDNGYSPFSFLSH.

[0070] The present invention has been described in detail above. Those skilled in the art will recognize that the invention can be practiced in a wide range of ways with equivalent parameters, concentrations, and conditions without departing from its spirit and scope, and without requiring unnecessary experiments. While specific embodiments have been provided, it should be understood that further modifications can be made to the invention. In summary, according to the principles of the invention, this application is intended to include any changes, uses, or improvements to the invention, including changes made using conventional techniques known in the art that depart from the scope disclosed herein.

Claims

1. The application of a protein or a biomaterial related to said protein, characterized in that, The application is any one of the following: E1) Regulates plant spike-type traits; E2) Preparation of products that regulate plant spikelet traits; E3) Plant breeding; The biological material is a substance that regulates the expression of the gene encoding the protein or a substance that regulates the content of the protein. The protein is any one of the following: B1) The amino acid sequence is that of the protein shown in SEQ ID NO:2 and / or SEQ ID NO:4; B2) A protein obtained by substituting and / or deleting and / or adding amino acid residues of the protein described in B1) that has more than 80% identity with and the same function as the protein shown in B1). B3) A fusion protein obtained by attaching a protein tag to the N-terminus and / or C-terminus of B1) or B2).

2. The application according to claim 1, characterized in that, The protein is derived from wheat.

3. The application according to any one of claims 1 or 2, characterized in that, The controlled plant spike-type trait is any one of the following: e1) Number of spikelets in the plant; e2) Number of plant florets.

4. The application according to any one of claims 1-3, characterized in that, The biomaterial is any one of the following: C1) Nucleic acid molecules that inhibit, reduce, or downregulate the expression of the gene encoding the protein described in claim 1 or 2; C2) expresses the gene encoding the nucleic acid molecule described in C1); C3) contains an expression cassette containing the gene described in C2); C4) A recombinant vector containing the gene described in C2), or a recombinant vector containing the expression cassette described in C3); C5) Recombinant microorganisms containing the gene described in C2), or recombinant microorganisms containing the expression cassette described in C3), or recombinant microorganisms containing the recombinant vector described in C4); C6) A transgenic plant cell line containing the gene described in C2), or a transgenic plant cell line containing the expression cassette described in C3), or a transgenic plant cell line containing the recombinant vector described in C4); C7) Transgenic plant tissue containing the gene described in C2), or transgenic plant tissue containing the expression cassette described in C3), or transgenic plant tissue containing the recombinant vector described in C4); C8) A transgenic plant organ containing the gene described in C2), or a transgenic plant organ containing the expression cassette described in C3), or a transgenic plant organ containing the recombinant vector described in C4).

5. The application according to claim 4, characterized in that, B1) The nucleic acid molecule is a gRNA that targets the protein-coding gene described in claim 1.

6. A method for regulating spike-type traits in plants, characterized in that, The method includes the following steps: the method includes introducing a substance into the target plant that inhibits, reduces or downregulates the expression of the gene encoding the protein described in claim 1 or 2, or inhibits, reduces or downregulates the activity and / or content of the protein, thereby altering the spike-like trait of the target plant, wherein the target plant contains the gene encoding the protein described in claim 1 or 2.

7. A method for cultivating plants with altered spikelet type traits, characterized in that, The method includes the following steps: the method includes introducing a substance into a target plant that inhibits, reduces, or downregulates the expression of the gene encoding the protein described in claim 1 or 2, or inhibits, reduces, or downregulates the activity and / or content of the protein, to obtain a plant with altered spikelet trait, wherein the target plant contains the gene encoding the protein described in claim 1 or 2.

8. The method according to claim 6 or 7, characterized in that, The introduction of a substance into the target plant that inhibits, reduces, or downregulates the expression of the gene encoding the protein described in claim 1 or 2, or inhibits, reduces, or downregulates the activity and / or content of the protein, includes introducing a gene knockout vector into the target plant targeting nucleotides 381 to 402 of SEQ ID NO:1 and nucleotides 390 to 411 of SEQ ID NO:3, or nucleotides 512 to 533 of SEQ ID NO:1 and nucleotides 521 to 542 of SEQ ID NO:

3.

9. The method according to any one of claims 6-8, characterized in that, The controlled plant spike-type trait is any one of the following: e1) Number of spikelets in the plant; e2) Number of plant florets.

10. The application according to any one of claims 1-5 or the method according to any one of claims 6-9, characterized in that, The plant is any one of the following: G1) Dicotyledons or monocotyledons; G2) Plants of the order Poales; G3) Gramineae plants; G4) Plants of the Triticum genus; G5) Wheat.

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