Application of wheat-derived protein and related biological materials thereof in improving yield per plant and grain protein content of crops

Abstract

The invention discloses application of protein derived from wheat and related biological materials of the protein in improving the yield of a single plant of a crop and the protein content of grains. According to the technical scheme protected by the invention, the protein derived from wheat is applied to improvement of the yield of a single plant and the protein content of plant grains, the name of the protein is SWEET11, and the protein can be a protein of which the amino acid sequence is a sequence 9, a sequence 6 or a sequence 3 in a sequence table. After the wheat SWEET11 protein is over-expressed in the national examination Zheng wheat 7698, the over-expressed TaSWEET11 is found to be capable of obviously improving the thousand seed weight of wheat grains and increasing the yield; meanwhile, overexpression of the TaSWEET11 enables the protein content of wheat grains to be increased, the quality of the grains is remarkably improved while the yield is increased, and the TaSWEET11 protein can be applied to crop breeding and quality improvement.

Description

Technical Field

[0001] The present invention belongs to the field of biotechnology, and particularly relates to the application of proteins derived from wheat and related biological materials thereof in increasing the yield per plant and the protein content of grains of crops. Background Art

[0002] Wheat is one of the important food crops in the world. With the continuous growth of the global population, the continuous reduction of arable land area, and the frequent occurrence of extreme weather, the biological and abiotic stresses during the whole growth period of wheat are aggravated, which poses a huge pressure on the improvement of wheat yield. With the continuous update of technology, the effective improvement of crop varieties has achieved a substantial increase in yield. In recent years, the increase in yield has reached a bottleneck period to a certain extent, and new strategies and ways to increase wheat yield need to be explored. The wheat yield is composed of three factors: the number of spikes per unit area, the number of grains per spike, and the 1000-grain weight. Therefore, increasing the 1000-grain weight is of great significance for improving the yield. Grain filling is the key process determining the grain yield of wheat. During the grain filling stage, the carbon source produced by the photosynthetic organs is transported over a long distance through the phloem to the grains, and storage substances such as starch are synthesized in the endosperm. At present, the mechanism of action of grain filling in wheat is not fully understood, and studying the regulation mechanism of grain development is of great significance for realizing high-yield breeding of wheat. Summary of the Invention

[0003] The technical problem to be solved by the present invention is how to increase the spike length, the number of spikelets and / or the number of grains per spike of a plant and / or how to increase the yield of wheat and improve the protein content of grains and / or how to increase the grain length, grain width and / or 1000-grain weight of plant grains and / or how to increase the yield per plant of a plant.

[0004] To solve the above technical problems, the present invention first provides any one of the following applications of a protein or a substance regulating protein expression or a substance regulating protein activity:

[0005] P1. Application in regulating the grain length, grain width and / or 1000-grain weight of plant grains,

[0006] P2. Application in increasing the grain length, grain width and / or 1000-grain weight of plant grains,

[0007] P3. Application in regulating the spike length, the number of spikelets / or the number of grains per spike of a plant,

[0008] P4. Application in increasing the spike length, the number of spikelets / or the number of grains per spike of a plant,

[0009] P5. Application in regulating the yield per plant of a plant,

[0010] P6. Application in increasing the yield per plant of a plant,

[0011] P7. Application in regulating the protein content of plant grains,

[0012] P8. Use in increasing the protein content in plant seeds

[0013] P9. Use in high-yield plant breeding and / or high-seed-protein-content breeding

[0014] P10. Use in improving the quality of high-yield plants and / or plants with high seed protein content

[0015] The protein may be the following proteins:

[0016] A1) A protein with an amino acid sequence that is Sequence 9 in the Sequence Listing;

[0017] A2) A protein with an amino acid sequence that is Sequence 3 in the Sequence Listing;

[0018] A3) A protein with an amino acid sequence that is Sequence 6 in the Sequence Listing;

[0019] A4) A protein obtained by substitution and / or deletion and / or addition of amino acid residues in the amino acid sequence shown in A1), A2) or A3), and having the same function, which is derived from A1), A2) or A3), or has more than 80% identity with the protein shown in A1), A2) or A3) and has the same function;

[0020] A4) A fusion protein obtained by connecting a protein tag to the N-terminus and / or C-terminus of A1), A2) or A3).

[0021] In the above applications, the protein may be derived from wheat.

[0022] The above protein can be artificially synthesized, or its coding gene can be synthesized first and then expressed biologically.

[0023] In the above protein, the protein tag refers to a polypeptide or protein that is fused and expressed with the target protein by using in vitro DNA recombination technology for the purpose of facilitating the expression, detection, tracing and / or purification of the target protein. The protein tag may be a Flag tag, His tag, MBP tag, HA tag, myc tag, GST tag and / or SUMO tag, etc.

[0024] For the above-mentioned proteins, 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 web page on the NCBI homepage website. For example, in Advanced BLAST 2.1, by 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 a search to calculate the identity of a pair of amino acid sequences, and then the identity value (%) can be obtained.

[0025] For the above-mentioned proteins, the identity of more than 80% may be at least 81%, 82%, 85%, 86%, 88%, 90%, 91%, 92%, 95%, 96%, 98%, 99%, or 100% identity.

[0026] In the above application, the plant may be any one of the following:

[0027] D1) Dicotyledonous plants;

[0028] D2) Monocotyledonous plants,

[0029] D3) Plants of the order Poales,

[0030] D4) Gramineous plants,

[0031] D5) Plants of the genus Triticum,

[0032] D6) Wheat.

[0033] To solve the above technical problems, the present invention also provides any one of the following applications of biomaterials related to the proteins described above:

[0034] Q1. Application in regulating the grain length, grain width, and / or 1000-grain weight of plant seeds,

[0035] Q2. Application in increasing the grain length, grain width, and / or 1000-grain weight of plant seeds,

[0036] Q3. Application in regulating the spike length, number of spikelets, and / or number of grains per spike of plant plants,

[0037] Q4. Application in increasing the spike length, number of spikelets, and / or number of grains per spike of plant plants,

[0038] Q5. Application in regulating the yield per plant of plant plants,

[0039] Q6. Application in increasing the yield per plant of plant plants,

[0040] Q7. Application in regulating the protein content in plant seeds,

[0041] Q8. Application in increasing the protein content in plant seeds,

[0042] Q9. Application in high-yield breeding of plants and / or breeding of plants with high seed protein content,

[0043] Q10. Application in improving the quality of high-yield plants and / or plants with high seed protein content,

[0044] The biological material is any one of the following B1) to B9):

[0045] B1) A nucleic acid molecule encoding the protein described in claim 1;

[0046] B2) An expression cassette containing the nucleic acid molecule described in B1);

[0047] B3) A recombinant vector containing the nucleic acid molecule described in B1), or a recombinant vector containing the expression cassette described in B2);

[0048] 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);

[0049] B5) A transgenic plant cell line containing the nucleic acid molecule described in B1), or a transgenic plant cell line containing the expression cassette described in B2);

[0050] B6) A transgenic plant tissue containing the nucleic acid molecule described in B1), or a transgenic plant tissue containing the expression cassette described in B2);

[0051] B7) A transgenic plant organ containing the nucleic acid molecule described in B1), or a transgenic plant organ containing the expression cassette described in B2);

[0052] B8) A nucleic acid molecule that promotes or increases the gene expression of the protein described in claim 1;

[0053] B9) An expression cassette, recombinant vector, recombinant microorganism or transgenic plant cell line containing the nucleic acid molecule described in B8).

[0054] In the above applications, the nucleic acid molecule described in B1) may be the coding gene of the protein shown in any of the following b1), b2) or b3):

[0055] b1) The coding sequence of the coding strand is the cDNA molecule or DNA molecule of sequence 8, sequence 2 and sequence 5 in the sequence listing;

[0056] b2) The nucleotide is the DNA molecule of sequence 7, sequence 1 and sequence 4 in the sequence listing,

[0057] b3) A cDNA molecule or a DNA molecule that hybridizes with the cDNA or DNA molecule defined in b2) and encodes a protein with the same function.

[0058] In the above applications, the plant can be any of the following:

[0059] D1) Dicotyledonous plants;

[0060] D2) Monocotyledonous plants,

[0061] D3) Plants of the order Poales,

[0062] D4) Gramineous plants,

[0063] D5) Plants of the genus Triticum,

[0064] D6) Wheat.

[0065] Among the above biological materials, the expression cassette containing a nucleic acid molecule described in B2) refers to DNA that can express the protein described in the above applications in a host cell. This DNA can not only include a promoter that initiates the transcription of the protein-coding gene, but also include a terminator that terminates the transcription of the protein-coding gene. Further, the expression cassette may also include enhancer sequences. Promoters that can be used in the present invention include, but are not limited to: constitutive promoters, tissue-, organ- and development-specific promoters, and inducible promoters.

[0066] An existing plant expression vector can be used to construct a recombinant expression vector containing the expression cassette of the protein-coding gene. The plant expression vectors include binary Agrobacterium vectors and vectors that can be used for plant microprojectile bombardment, etc. Such as pAHC25, pWMB123, pBin438, pCAMBIA1302, pCAMBIA2301, pCAMBIA1301, pCAMBIA1300, pBI121, pCAMBIA1391-Xa or pCAMBIA1391-Xb (from CAMBIA), etc. The plant expression vector may also contain the 3'-untranslated region of the foreign gene, that is, it contains a polyadenylation signal and any other DNA fragments 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. For example, the non-translated regions transcribed at the 3' end of the Agrobacterium tumefaciens Ti plasmid gene (such as the nopaline synthase gene Nos) and plant genes (such as the soybean storage protein gene) have similar functions. When using the gene of the present invention to construct a plant expression vector, enhancers can also be used, including translational enhancers or transcriptional enhancers. These enhancer regions can be the ATG start codon or the start codon in the adjacent region, etc., but must have the same reading frame as the coding sequence to ensure the correct translation of the entire sequence.

[0067] Among the above-mentioned biological materials, the recombinant microorganism can specifically be yeast, bacteria, algae and fungi.

[0068] To solve the above technical problems, the present invention also provides a method for increasing the grain length, grain width, thousand-grain weight, number of grains per ear, yield per plant and grain protein content of plants, including enhancing or increasing the activity of the protein described above in the target plant and / or the expression level of the coding gene of the protein described above, thereby increasing the grain length, grain width, thousand-grain weight, number of grains per ear, yield per plant and grain protein content of the target plant.

[0069] In the above method, the enhancing or increasing the activity of the protein described above in the target plant and / or the expression level of the coding gene of the protein described above is achieved by introducing the coding gene of the protein described above into the target plant.

[0070] In the above method, the coding gene of the protein can be first modified as follows and then introduced into the target plant to achieve a better expression effect:

[0071] 1) Connect with promoters expressed in various plants to facilitate its expression in plants; the promoters can include constitutive, inducible, temporal regulation, developmental regulation, chemical regulation, tissue-preferred and tissue-specific promoters; the selection of promoters will vary with the time and space requirements of expression and also depends on the target species; for example, tissue or organ-specific expression promoters are determined according to the developmental stage of the receptor as needed; although many promoters derived from dicotyledonous plants have been proven to be functional in monocotyledonous plants and vice versa, ideally, dicotyledonous plant promoters are selected for expression in dicotyledonous plants and monocotyledonous plant promoters are selected for expression in monocotyledonous plants;

[0072] 2) Connect with suitable transcription terminators, which can also improve the expression efficiency of the gene of the present invention; for example, tml derived from CaMV and E9 derived from rbcS; any available terminator known to be functional in plants can be connected to the gene of the present invention;

[0073] 3) Introduce enhancer sequences, such as intron sequences (for example, derived from Adhl and bronzel) and viral leader sequences (for example, derived from TMV, MCMV and AMV).

[0074] In the above method, the stress-sensitive plants can be transgenic plants or plants obtained by conventional breeding techniques such as hybridization.

[0075] In the above method, the transgenic plants are understood to include not only the first-generation to second-generation transgenic plants, but also their progeny. For transgenic plants, the gene can be propagated in the species, or transferred into other varieties of the same species, particularly including commercial varieties, using conventional breeding techniques. The transgenic plants include seeds, callus, whole plants, and cells.

[0076] In the above method, the plant and / or the target plant can be any one of the following:

[0077] D1) Dicotyledonous plants;

[0078] D2) Monocotyledonous plants,

[0079] D3) Plants of the order Poales,

[0080] D4) Gramineous plants,

[0081] D5) Plants of the genus Triticum,

[0082] D6) Triticum aestivum.

[0083] The proteins and / or biomaterials described above also fall within the scope of protection of the present invention.

[0084] In the present invention, the gene of wheat SWEET11 protein is edited and knocked out, which can significantly reduce the plant yield; further overexpression of this gene reveals that the thousand-grain weight of the overexpression material increases, the protein content is improved, and the yield per plant is increased. The present invention has important theoretical and practical significance for improving wheat yield through genetic engineering techniques and methods.

[0085] With the increase in the global population, improving yield remains an urgent goal in wheat breeding. In the past few decades, the thousand-grain weight of grains, the number of grains per spike, and the tiller number have been issues of great concern to breeders. Research on improving wheat yield by genetically modifying the expression characteristics of grain weight genes has been widely reported. The transport of sugars is a prerequisite for the synthesis of starch in the developing endosperm of grains, and the efficient transport of sucrose is the material basis for grain filling. The loss of function of sugar transporters will lead to defects in grain filling and the inability to form a functional endosperm, thus significantly reducing grain yield. BRIEF DESCRIPTION OF THE DRAWINGS

[0086] Figure 1Phenotypic comparisons of plant height, spike length, yield per plant, and grain morphology between wild-type Fielder (WT) and its TaSWEET11 gene-edited (KO) materials. A shows the comparison of plant height between wild-type materials and TaSWEET11 gene-edited materials, with the vertical axis representing plant height; B shows the comparison of spike length between wild-type materials and TaSWEET11 gene-edited materials, with the vertical axis representing spike length; C shows the comparison of the number of spikelets between wild-type materials and TaSWEET11 gene-edited materials, with the vertical axis representing the number of spikelets per plant; D shows the comparison of the number of grains per spike between wild-type materials and TaSWEET11 gene-edited materials, with the vertical axis representing the number of grains per spike; E shows the comparison of 1000-grain weight between wild-type materials and TaSWEET11 gene-edited materials, with the vertical axis representing 1000-grain weight; F shows the comparison of grain length between wild-type materials and TaSWEET11 gene-edited materials, with the vertical axis representing grain length; G shows the comparison of grain width between wild-type materials and TaSWEET11 gene-edited materials, with the vertical axis representing grain width; H shows the comparison of yield per plant between wild-type materials and TaSWEET11 gene-edited materials, with the vertical axis representing yield per plant. I shows a photo of the comparison of grain length, grain width, and grain morphological size between wild-type materials and TaSWEET11 gene-edited materials.

[0087] Figure 2 Phenotypic comparisons of plant height, spike length, and yield per plant between ZM7698 and its TaSWEET11 overexpressed (OE) materials. A shows the comparison of plant height between ZM7698 and its TaSWEET11 overexpressed (OE) materials, with the vertical axis representing plant height; B shows the comparison of spike length between ZM7698 and its TaSWEET11 overexpressed (OE) materials, with the vertical axis representing spike length; C shows the comparison of the number of spikelets between ZM7698 and its TaSWEET11 overexpressed (OE) materials, with the vertical axis representing the number of spikelets per plant; D shows the comparison of the number of grains per spike between ZM7698 and its TaSWEET11 overexpressed (OE) materials, with the vertical axis representing the number of grains per spike; E shows the comparison of grain length between ZM7698 and its TaSWEET11 overexpressed (OE) materials, with the vertical axis representing grain length; F shows the comparison of grain width between ZM7698 and its TaSWEET11 overexpressed (OE) materials, with the vertical axis representing grain width; G shows the comparison of 1000-grain weight between ZM7698 and its TaSWEET11 overexpressed (OE) materials, with the vertical axis representing 1000-grain weight; H shows the comparison of yield per plant between ZM7698 and its TaSWEET11 overexpressed (OE) materials, with the vertical axis representing yield per plant. I shows a photo of the comparison of grain length and grain width between ZM7698 and its TaSWEET11 overexpressed (OE) materials.

[0088] Figure 3 Analysis and comparison of grain protein content between wild-type wheat ZM7698 and TaSWEET11 overexpressed (OE) materials. The vertical axis represents the grain protein content (%).

[0089] Figure 4PCR amplification identification of the SWEET11 gene in wild-type wheat ZM7698 and TaSWEET11 overexpression OE materials.

[0090] Figure 5 Detection of the expression level of the TaSWEET11 gene in wild-type wheat ZM7698 and TaSWEET11 overexpression OE materials Detailed implementation mode

[0091] The present invention will be further described in detail below in conjunction with the detailed implementation mode. The provided embodiments are only for clarifying the present invention, rather than limiting the scope of the present invention. The following provided embodiments can be used as a guide for those of ordinary skill in the art to make further improvements, and do not limit the present invention in any way.

[0092] The experimental methods in the following embodiments, unless otherwise specified, are all conventional methods, and are carried out according to the techniques or conditions described in the literature in this field or according to the product instructions. The materials, reagents, etc. used in the following embodiments, unless otherwise specified, can all be obtained from commercial channels.

[0093] Example 1: CRISPR target site search and vector construction

[0094] 1. Target site and primer design

[0095] The wheat SWEET11 protein includes three homologous proteins, TaSWEET11-7A, TaSWEET11-7B, and TaSWEET11-7D, which are distributed on the A, B, and D sub-genome chromosomes of wheat, respectively. The genomic nucleotide sequence of the wheat TaSWEET11-7A gene is shown as Sequence 1 in the sequence listing, the CDS sequence of the TaSWEET11-7A gene is shown as Sequence 2 in the sequence listing, and the amino acid sequence of the TaSWEET11-7A protein is shown as Sequence 3 in the sequence listing; the genomic nucleotide sequence of the wheat TaSWEET11-7B gene is shown as Sequence 4 in the sequence listing, the CDS sequence of the TaSWEET11-7B gene is shown as Sequence 5 in the sequence listing, and the amino acid sequence of the TaSWEET11-7B protein is shown as Sequence 6 in the sequence listing; the genomic nucleotide sequence of the wheat TaSWEET11-7D gene is shown as Sequence 7 in the sequence listing, the CDS sequence of the TaSWEET11-7D gene is shown as Sequence 8 in the sequence listing, and the amino acid sequence of the TaSWEET11-7D protein is shown as Sequence 9 in the sequence listing.

[0096] The sequence of Sequence 1 is as follows (5'-3'):

[0097]

[0098] The sequence of Sequence 2 is as follows (5'-3'):

[0099] ATGGCTGGGGGCCTCTTCGACATGTCTCACCCGGCGAGCGCCCTCGCCGGCATCGCAGGAAACATCGTCTCCTTCTTCGTGTTCCTCGCGCCAATGGCGACGTTCCTGCAGATCTACAGGAAGAAGACGACGGGCGGGTTCAGCTCGGTGCCGTACGTGGTGGCGCTCTTCAGCTGCTCGCTGCTCATCTTCTACGCGCTGCTCAAGACGAAGTCCCCCCTCCTGCTCACCATCAACAGCTTCGGCTGCTGCATCGAGACCGTCTACATCATCGCCTACCTCGTCTACGCGCCGCCCCGCGCCAGGCTCAGGACGCTCGCCTACTTCTTCGTCCTCGACGTCGCCGCCTTCGGCCTCGTCCTCGTCGTCACCATGTACGCCTTCGCGCCGGCCCACCGCGTCAAGTTCCTCGGCAGCGTCTGCCTCGCCTTCTCCATGGCCGTCTTCGTCGCGCCGCTCTCCATCATCGTCAAGGTGATCAAGACCAAGAGCGTGGAGTTCCTGCCCGTCGGCCTCTCCTTCTGCCTCGTCCTCAGCGCCGTCGCGTGGTTCTGCTACGGCCTCTTCACCAAGGACCCATTCGTCATGTACCCAAACGTGGGCGGCTTCTTCTTCAGCTGCGTCCAGATCGGGCTCTACTGCTGGTACCGCAAGCCCAGCAACGCCGTGCTGCCCACTACCACCGCCGACGCCGGCAACGGCAATGGCGGCGCCGCGGAGCAGCCGCAGGTGCAGGTGATCGAGCTGCCCGCGCACTCCGTCGCCATCCTGTCCGTGGGCCCCGTCCCGATCCTCGGCGTGCACAAGATCGAGGTCATGGCGGCAGAGCAGCAGACCGTCATCGACGTCAAGGACGCCGCCGAGGCCGACCAGCCGGAGGTCATCGAGATCGTCCCCGCCCCCGCCGTGTGA。

[0100] The sequence of Sequence 3 is as follows:

[0101] MAGGLFDMSHPASALAGIAGNIVSFFVFLAPMATFLQIYRKKTTGGFSSVPYVVALFSCSLLIFYALLKTKSPLLLTINSFGCCIETVYIIAYLVYAPPRARLRTLAYFFVLDVAAFGLVLVVTMYAFAPAHRVKFLGSVCLAFSMAVFVAPLSIIVKVIKTKSVEFLPVGLSFCLVLSAVAWFCYGLFTKDPFVMYPNVGGFFFSCVQIGLYCWYRKPSNAVLPTTTADAGNGNGGAAEQPQVQVIELPAHSVAILSVGPVPILGVHKIEVMAAEQQTVIDVKDAAEADQPEVIEIVPAPAV.

[0102] The sequence of Sequence 4 (5'-3') is as follows:

[0103]

[0104] The sequence of Sequence 5 is as follows (5’-3’):

[0105] ATGGCTGGGGGCCTCTTCGACATGTCTCACCCGGCGAGCGCCCTCGCCGGCATCGCAGGAAACATCGTCTCCTTCTTCGTGTTCCTCGCGCCAATGGCGACGTTCCTGCAGATCTACAGGAAGAAGACGACGGGCGGGTTCAGCTCGGTGCCGTACGTGGTGGCGCTCTTCAGCTGCTCGCTGCTCATCTTCTACGCGCTGCTCAAGACCCACTCCCCCCTCCTGCTCACCATCAACAGCTTCGGCTGCTGCATCGAGACCGTCTACATCATCGCCTACCTCGTCTACGCGCCGCCCCGGGCCAGGCTCAGGACGCTCGCCTACTTCTTCGTCCTCGACGTCGCCGCCTTCGGCCTCGTCCTCGTCGTCACCATGTACGCCTTCGCGCCGGCCCACCGCGTCAAGTTCCTCGGCAGCGTCTGCCTCGCCTTCTCCATGGCCGTCTTCGTCGCGCCGCTCTCCATCATCGTCAAGGTGATCAAGACCAAGAGCGTGGAGTTCCTGCCCGTCGGCCTCTCCTTCTGCCTCGTCCTCAGCGCTGTCGCCTGGTTCTGCTACGGCCTCTTCACCAAGGACCCATTCGTCATGTACCCGAACGTGGGCGGCTTCTTCTTCAGCTGCGTCCAGATTGGGCTCTACTGCTGGTACCGCAAGCCCAGCAACGCCGTGCTGCCCACTACTACCGCCGACGCCGGCAACGGCAATGGCGGCTCCGCGGACCAGCCGCAGGTGCAGGTGATCGAGCTGCCCGTGCACTCCGTCGCCATCCTGTCCGTGGGCCCCGTCCCGATCCTCGGCGTGCACAAGATCGAGGTCATCGCAGCAGAGCAGCAGACCGTCATCGACGTCAAGGACGCCGCCAAGGCGGCCGAGGTCGACCAGCCGGAGGTCATCGAGATCGTCCCCGCCCCCGCCGTGTGA。

[0106] The sequence of Sequence 6 is as follows:

[0107] MAGGLFDMSHPASALAGIAGNIVSFFVFLAPMATFLQIYRKKTTGGFSSVPYVVALFSCSLLIFYALLKTHSPLLLTINSFGCCIETVYIIAYLVYAPPRARLRTLAYFFVLDVAAFGLVLVVTMYAFAPAHRVKFLGSVCLAFSMAVFVAPLSIIVKVIKTKSVEFLPVGLSFCLVLSAVAWFCYGLFTKDPFVMYPNVGGFFFSCVQIGLYCWYRKPSNAVLPTTTADAGNGNGGSADQPQVQVIELPVHSVAILSVGPVPILGVHKIEVIAAEQQTVIDVKDAAKAAEVDQPEVIEIVPAPAV.

[0108] The sequence of Sequence 7 (5’-3’) is as follows:

[0109]

[0110] The sequence of Sequence 8 is as follows (5'-3'):

[0111] ATGGCTGGGGGCCTCTTCGACATGTCTCACCCGGCGAGCGCCCTCGCCGGCATCGCAGGAAACATCGTCTCCTTCTTCGTGTTCCTCGCGCCAATGGCGACGTTCCTGCAGATCTACAGGAAGAAGACGACGGGCGGGTTCAGCTCGGTGCCGTACGTGGTGGCGCTCTTCAGCTGCTCGCTGCTCATCTTCTACGCGCTGCTCAAGACCGACTCCCCCCTCCTGCTCACCATCAACAGCTTCGGCTGCTGCATCGAGACCGTCTACATCGTCGCCTACCTCGTCTACGCGCCGCCCCGCGCCAGGCTCAGGACGCTCGCCTACTTCTTCGTCCTCGACGTCGCCGCCTTCGGCCTCGTCCTCGTCGTCACCATGTACGCCTTCGCGCCGGCCCACCGCGTCAAGTTCCTCGGCAGCGTCTGCCTCGCATTCTCCATGGCCGTCTTCGTCGCGCCGCTCTCCATCATCGTCAAGGTGATCAAGACCAAGAGCGTGGAGTTCCTGCCCGTCGGCCTCTCCTTCTGCCTCGTCCTCAGCGCCGTCGCGTGGTTCTGCTACGGCCTCTTCACCAAGGACCCATTCGTCATGTACCCGAACGTGGGCGGCTTCTTCTTCAGCTGCGTCCAGATTGGGCTCTACTGCTGGTACCGCAAGCCCAGCAACGCCGTGCTGCCCACTACTACCGCCGACGCCGGCAACGGCAATGGCGGCTCCGCAGACCAGCCGCAGGTGCAGGTGATCGAGCTGCCCGTGCACTCCGTCGCCATCCTGTCCGTGGGCCCCGTCCCGATCCTCGGCGTGCACAAGATCGAGGTCATCGCGGCAGAGCAGCAGACCGTCATCGACGTCAAGGACGCCGCCAGGGCTGCCGAGGTCGACCAGCCGGAGGTCATCGAGATCGTCCCCGCCCCCGCCGTGTGA。

[0112] The sequence of Sequence 9 is as follows:

[0113] MAGGLFDMSHPASALAGIAGNIVSFFVFLAPMATFLQIYRKKTTGGFSSVPYVVALFSCSLLIFYALLKTDSPLLLTINSFGCCIETVYIVAYLVYAPPRARLRTLAYFFVLDVAAFGLVLVVTMYAFAPAHRVKFLGSVCLAFSMAVFVAPLSIIVKVIKTKSVEFLPVGLSFCLVLSAVAWFCYGLFTKDPFVMYPNVGGFFFSCVQIGLYCWYRKPSNAVLPTTTADAGNGNGGSADQPQVQVIELPVHSVAILSVGPVPILGVHKIEVIAAEQQTVIDVKDAARAAEVDQPEVIEIVPAPAV。

[0114] Use the E-CRISP Design website (http: / / www.e-crisp.org / E-CRISP / designcrispr.html) to search for 2 gRNA target sites conserved in each sub-genome of wheat A, B, and D in the genomic sequence of wheat TaSWEET11 (gRNA1 sequence: 5’-gACGACGGGCGGGTTCAGCT-3’, gRNA2 sequence: 5’-TCCCGATCCTCGGCGTGCAC-3’). Use the Ensembl Plants website (http: / / plants.ensembl.org / index.html) to align the specificity of the target sites. Design primers (Primer-F: 5’-aataatggtctcAAGCgACGACGGGCGGGTTCAGCT-3’; Primer-F0: 5’-gACGACGGGCGGGTTCAGCTgttttagagctagaaatagc-3’; Primer-R: 5’-ATTATTGGTCTCTAAACTCCCGATCCTCGGCGTGCAC-3’; Primer-R0: 5’-TCCCGATCCTCGGCGTGCACCGCTTCTTGGTGCC-3’) to amplify plasmid pCBC-MT1T2 (preserved in this laboratory, related literature: Du D, Zhang D, Yuan J, Feng M, Li Z, Wang Z, Zhang Z, Li X, Ke W, Li R, Chen Z, Chai L, Hu Z, Guo W, Xing J, Su Z, Peng H, Xin M, Yao Y, Sun Q, Liu J, Ni Z. FRIZZY PANICLE defines a regulatory hub for simultaneously controlling spikelet formation and awn elongation in bread wheat. New Phytol. 2021 Jul;231(2):814-833. The public can obtain it from the applicant and is only used to repeat this invention).

[0115] 2. PCR Amplification and Product Purification and Recovery

[0116] For PCR amplification, use Tks-DNA polymerase (Takara, R060A), and the reaction system is as follows:

[0117]

[0118] The PCR reaction program was set as follows: pre-denaturation at 94°C for 5 min; denaturation at 94°C for 10 s, annealing at 58°C for 15 s, extension at 68°C for 30 s, for a total of 35 cycles; extension at 68°C for 5 min, and storage at 4°C.

[0119] Agarose gel recovery: Loading Buffer was added to the PCR reaction product, and then agarose gel electrophoresis was performed at 180 V for 10 min. After electrophoresis, the target band was cut and recovered under ultraviolet light. The steps are as follows:

[0120] 1) The cut gel block was placed into a 2.0 mL optimized centrifuge tube, 300 μL of solubilization solution was added, and it was incubated in a 55°C water bath for 10 - 15 min, and inverted up and down 1 - 2 times until the gel block was completely dissolved;

[0121] 2) After it cooled to room temperature, the adsorption column was placed in the collection tube, the solubilization mixture was aspirated into the adsorption column, allowed to stand for 3 min, and centrifuged at 12,000 rpm for 1 min;

[0122] 3) 500 μL of wash buffer W1 was added to the adsorption column, and centrifuged at 12,000 rpm for 1 min;

[0123] 4) 700 μL of wash buffer W2 was added, allowed to stand for 1 min, and centrifuged at 12,000 rpm for 1 min; repeated once;

[0124] 5) Left at room temperature for 5 - 10 min, and the adsorption column was placed into a new 1.5 mL centrifuge tube;

[0125] 6) 20 - 30 μL of ddH2O was added, left at room temperature for 2 min, and centrifuged at 12,000×rpm for 1 min. The concentration was measured and stored at -20°C.

[0126] 3. Construction of CRISPR vector

[0127] The recovered product of glue obtained in step 2 and the vector pBUE411 (preserved in our laboratory, related literature: Du D, Zhang D, Yuan J, Feng M, Li Z, Wang Z, Zhang Z, Li X, Ke W, Li R, Chen Z, Chai L, Hu Z, Guo W, Xing J, Su Z, Peng H, Xin M, Yao Y, Sun Q, Liu J, Ni Z. FRIZZY PANICLE defines a regulatory hub for simultaneously controlling spikelet formation and awn elongation in bread wheat. New Phytol. 2021 Jul; 231(2):814-833. The public can obtain it from the applicant and is only used to repeat the present invention) are reacted through the following system:

[0128]

[0129] Reaction procedure: 37 °C, 5 h; 50 °C, 5 min; 80 °C, 10 min.

[0130] After the reaction product is transformed into Escherichia coli competent cells, the transformed bacteria are detected by PCR amplification using primers 411-F (5’-TTTCCCAGTCACGACGTTGT-3’) and 411-R (5’-GGATTCATGAGCAGCAAGCA-3’). The transformed bacteria containing the target sequences of gRNA1 and gRNA2 are positive monoclonal bacteria. The positive monoclonal bacteria are inoculated into LB liquid medium, and then the plasmid is extracted from the cultured bacterial solution to obtain the CRISPR-Cas9 recombinant plasmid pBUE411-TaSWEET11.

[0131] Example 2, Gene amplification of TaSWEET11 and construction of overexpression vector

[0132] 1. Amplification of target gene

[0133] Using the cDNA of wheat variety XX329 as a template (preserved in our laboratory, related literature: Zhang H, Zhu B, Qi B, Gou X, Dong Y, Xu C, Zhang B, Huang W, Liu C, Wang X, Yang C, Zhou H, Kashkush K, Feldman M, Wendel JF, Liu B. Evolution of the BBAA component of bread wheat during its history at the allohexaploid level. Plant Cell. 2014 Jul;26(7):2761 - 76. The public can obtain it from the applicant and is only used to repeat the present invention), PCR amplification was carried out using Tks - DNA polymerase to obtain the CDS sequence of the TaSWEET11 - 7D gene. The primer sequences are as follows:

[0134] Primer - F1: 5’ - ACACTAGTTCTTGGCTAA - 3’;

[0135] Primer - R1: 5’ - GTCGTCGTCGATGTTGGTG - 3’.

[0136] The reaction system is as follows:

[0137]

[0138] The PCR reaction program was set as: pre - denaturation at 94°C for 5 min; denaturation at 94°C for 10 s, annealing at 58°C for 15 s, extension at 68°C for 30 s, for a total of 35 cycles; extension at 68°C for 5 min, and stored at 4°C.

[0139] 2. Purification and recovery of PCR products

[0140] The PCR amplification products obtained in step 1 were purified and recovered to obtain purified and recovered products. The purification and recovery steps of PCR products were the same as those in step 2 of Example 1.

[0141] 3. Construction of TaSWEET11 gene over - expression vector:

[0142] 3.1 Linearization of the expression vector

[0143] According to the multiple cloning site information of vector pWMB110 (preserved in our laboratory, related literature: Liu J, Chen Z, Wang Z, Zhang Z, Xie X, Wang Z, Chai L, Song L, Cheng X, Feng M, Wang X, Liu Y, Hu Z, Xing J, Su Z, Peng H, Xin M, Yao Y, Guo W, Sun Q, Liu J, Ni Z. Ectopic expression of VRT-A2 underlies the origin of Triticum polonicum and Triticum petropavlovskyi with long outer glumes and grains. Mol Plant. 2021 Sep 6;14(9):1472-1488. The public can obtain it from the applicant and is only used to repeat the present invention), the vector pWMB110 was digested with the restriction endonuclease BamHI to obtain a linearized vector plasmid.

[0144] 3.2 PCR amplification

[0145] Using the recovered product in step 2 as a template, primers ubi-SWT-F: 5’-AGGTCGACTCTAGAGGATCCATGGCTGGGGGCCTCTTC-3’ and ubi-SWT-R: 5’-AGCTCGGTACCCGGGGATCCTCACACGGCGGGGGCGGGGA-3’ were used for PCR amplification. The amplification system and procedure were the same as those for gene amplification in step 1. The vector homologous sequences were ligated to both ends of the TaSWEET11-7D sequence to obtain a PCR product. The same method as in step 2 was used to recover this PCR product to obtain a PCR purified and recovered product, which was the TaSWEET11-7D target sequence containing homologous sequences at both ends of the vector.

[0146] 4. Seamless cloning ligation reaction

[0147] The ligation system was as follows: 2×Seamless Cloning Mix (Bomed, CL117-01), 2.5 μL; the linearized pWMB110 vector obtained in step 3.1, 1.5 μL; the PCR purified and recovered product obtained in step 3.2, 1 μL. React at 50 °C for 30 min to obtain the recombinant plasmid pWMB110-Ubi-TaSWEET11.

[0148] 5. Obtaining positive recombinant bacteria

[0149] The recombinant plasmid pWMB110-Ubi-TaSWEET11 obtained in step 4 was transformed into Escherichia coli competent cells to obtain recombinant Escherichia coli.

[0150] PCR was used to detect the recombinant Escherichia coli. The detection primers were: Ubi-F: 5’-TCGATGCTCACCCTGTTGTTTG-3’; OS-R: 5’-GGACGATCTCGATGACCT-3’. The bacterial solution containing the target fragment of about 700 bp in size (containing the CDS sequence of the TaSWEET11-7D gene shown in Sequence 8 in the Sequence Listing) in the PCR product was sent for sequencing. According to the sequencing results, the positive bacterial solution was retained to obtain positive recombinant Escherichia coli.

[0151] The experimental steps for transforming the ligation product into Escherichia coli competent cells are as follows:

[0152] 1) Place the Escherichia coli competent cells on ice. When they are in the freeze-thaw state, add the recombinant plasmid pWMB110-Ubi-TaSWEET11, gently mix well, and incubate on ice for 30 min.

[0153] 2) After that, perform a water bath at 42 °C for 1 min; then incubate on ice for 2 min, add 500 μL of LB liquid medium, and culture with shaking at 37 °C for 1 h.

[0154] 3) Centrifuge and retain 200 μL of the bacterial solution, spread it on the LB-resistant solid medium, and culture it upside down at 37 °C for 1 day.

[0155] 4) Pick a single clone into 1 mL of LB liquid medium, culture it with shaking at 37 °C, screen the positive bacterial solution by PCR, mix it with 50% glycerol (volume ratio 1:1), and store it at -80 °C for later use.

[0156] The positive recombinant Escherichia coli was cultured with shaking for expansion, and then the recombinant plasmid pWMB110-Ubi-TaSWEET11 was extracted using a plasmid extraction kit (Zhuangmeng, ZP101-1).

[0157] Example 3. Obtaining of TaSWEET11 recombinant Agrobacterium

[0158] The CRISPR-Cas9 recombinant plasmid pBUE411-TaSWEET11 obtained in Example 1 and the recombinant overexpression plasmid pWMB110-Ubi-TaSWEET11 obtained in Example 2 were respectively transformed into Agrobacterium. The experimental steps are as follows:

[0159] 1) When the Agrobacterium tumefaciens competent cell EHA105 (Vidi, AC1010) is in the freeze-thaw state, add a certain volume of the target plasmid (about 1 μg, pBUE411-TaSWEET11 or pWMB110-Ubi-TaSWEET11), gently mix, and place on ice for 5 min.

[0160] 2) Quickly place the competent cells in liquid nitrogen for 5 min and then in a 37 °C water bath for 5 min.

[0161] 3) Place on ice for 5 min, add 750 μL of LB liquid medium, and culture with shaking at 28 °C for 2 - 3 h.

[0162] 4) Pipette 200 μL of the bacterial solution and spread it on an LB solid medium containing Rif and Kan, and culture it upside down at 28 °C for 1 - 2 days.

[0163] Pick a monoclonal plaque, add 1 mL of LB liquid medium, and culture with shaking at 28 °C for 1 day to obtain the positive recombinant Agrobacterium EHA105 / pBUE411-TaSWEET11 bacterial solution and the positive recombinant Agrobacterium EHA105 / pWMB110-Ubi-TaSWEET11 bacterial solution. Mix them with 50% glycerol (volume ratio 1:1) and store at -80 °C for later use.

[0164] The positive recombinant Agrobacterium EHA105 / pWMB110-Ubi-TaSWEET11 contains the CDS sequence of the TaSWEET11-7D gene shown in Sequence 8 in the sequence listing and can express TaSWEET11-7D shown in Sequence 9 in the sequence listing.

[0165] Example 4. Positive identification of transgenic wheat materials and determination of agronomic traits

[0166] 1. Obtaining transgenic wheat materials

[0167] The recombinant Agrobacterium EHA105 / pBUE411-TaSWEET11 and recombinant Agrobacterium EHA105 / pWMB110-Ubi-TaSWEET11 obtained in Example 3 were respectively used to transform the callus of wheat by Agrobacterium-mediated method. Among them, the recombinant Agrobacterium EHA105 / PBUE411-TaSWEET11 containing the CRISPR vector was used to transform wheat Fielder to obtain the gene-edited material KO lines of TaSWEET11 in the T0 generation, and the genetic transformation was carried out by the transgenic platform of the Wheat Research Center of China Agricultural University; the recombinant Agrobacterium EHA105 / pWMB110-Ubi-TaSWEET11 was used to transform Guoshen Zhengmai 7698 (related literature: Luo J, Li S, Xu J, Yan L, Ma Y, Xia L. Pyramiding favorable alleles in an elite wheat variety in one generation by CRISPR-Cas9-mediated multiplex gene editing. Mol Plant. 2021 Jun 7;14(6):847-850. The public can obtain it from the applicant and is only used to repeat the present invention) to obtain the overexpression OE materials OE1, OE2 and OE3 of TaSWEET11 in the T0 generation. The genetic transformation of wheat callus was entrusted to the Research Group of Gene Editing and New Material Creation of the Institute of Crop Science, Chinese Academy of Agricultural Sciences (experimental procedures refer to the literature: Luo J, Li S, Xu J, Yan L, Ma Y, Xia L. Pyramiding favorable alleles in an elite wheat variety in one generation by CRISPR-Cas9-mediated multiplex gene editing. Mol Plant. 2021 Jun 7;14(6):847-850).

[0168] 2. Identification of gene-edited plants:

[0169] DNA of the gene-edited materials in the T0 generation was extracted, and the gene-edited materials in the T0 generation were subjected to PCR amplification using three groups of CRISPR detection primers, and the amplification products were sequenced. Plants with mutations in different chromosomal subgroups of ABD were screened, subcultured, and homozygous triple mutant plants were obtained for planting to obtain three T2-generation plants KO1, KO2 and KO3, and their agronomic traits were investigated.

[0170] The sequences of the three groups of CRISPR detection primers are as follows:

[0171] SWTCSA-F: 5'-AGTGGGTGGATCTCTGACTCC-3',

[0172] SWTCSA-R: 5'-TCGTGTCGTCGTCGATGTTT-3';

[0173] SWTCSB-F: 5'-CTGTCGCCTGGTTCTGCT-3',

[0174] SWTCSB-R: 5'-ACACCAACCTGCGGACTAATA-3';

[0175] SWTCSD-F: 5'-TCCATCATCGTACGTCGTTAGC-3',

[0176] SWTCSD-R: 5'-TATTGTCACCGCACCAACG-3'.

[0177] 3. Identification of positive overexpression plants:

[0178] Using the positive identification primers for OE materials (Ubi-F: 5'-TCGATGCTCACCCTGTTGTTTG-3'; OS-R: 5'-GGACGATCTCGATGACCT-3'), PCR amplification was performed on the DNA of the T0 generation of the TaSWEET11 gene overexpression OE materials to determine the positive lines ( Figure 4 ) After two generations of screening and planting, the T2 generation pure line plants OE6, OE8, and OE10 were obtained, and the expression levels of the TaSWEET11-7D gene were measured. The results are as Figure 5 shown. Compared with the wild-type receptor wheat Guoshen Zhengmai 7698 ( Figure 5 ZM7698 in it), the TaSWEET11-7D gene was significantly overexpressed and was subsequently used for phenotypic research.

[0179] 4. Measurement of agronomic traits:

[0180] At the mature stage of wheat, the agronomic traits such as plant height, spike length, number of spikelets, number of grains per spike, yield per plant, and grain length, grain width, and 1000-grain weight of the grains of the T2 generation TaSWEET11 gene-edited plant KO lines, the T2 generation overexpression material OE lines, and the wild-type materials were investigated; the crude protein content in the seeds was measured by the Kjeldahl method.

[0181] The identification results of the agronomic traits are as follows:

[0182] The agronomic traits of the KO materials (KO1, KO2, and KO3) were in plant height ( Figure 1 A in it), spike length ( Figure 1in B), the number of spikelets ( Figure 1 in C) and the number of grains per spike ( Figure 1 in D) were significantly reduced phenotypically compared with the wild-type receptor material ( Figure 1 in WT), and the grain length ( Figure 1 in F), grain width ( Figure 1 in G) and 1000-grain weight ( Figure 1 in E) of the grains were extremely significantly reduced, resulting in a serious reduction in the yield per plant ( Figure 1 in H). Figure 1 ).

[0183] By using the Ubi promoter to drive the heterologous overexpression of the TaSWEET11 gene in the background of the national-approved winter wheat variety Zhengmai 7698, it was found that the OE lines (OE6, OE8 and OE10) of Zhengmai 7698 compared with Zhengmai 7698 ( Figure 2 in ZM7698) had a reduced plant height ( Figure 2 in A), and the spike length ( Figure 2 in B), the number of spikelets ( Figure 2 in C) and the number of grains per spike ( Figure 2 in D) all increased. The grain length ( Figure 2 in E) and grain width ( Figure 2 in F) of the grains of the OE material both increased significantly, increasing the 1000-grain weight ( Figure 2 in G) by about 1.50 - 1.82 g; the yield per plant increased significantly, with an increase range of about 20% - 49% ( Figure 2 in H). By analyzing the protein content of the grains, it was found that the content of the grains of the OE material was significantly increased compared with Zhengmai 7698 ( Figure 3 in ZM7698) ( Figure 3 ).

[0184] Therefore, overexpressing TaSWEET11 can significantly increase the 1000-grain weight of wheat grains and improve the yield; at the same time, overexpressing TaSWEET11 also increases the protein content of wheat grains, significantly improving the grain quality while increasing the yield. The wheat TaSWEET11 protein can be applied to crop breeding and quality improvement.

[0185] The above has described the present invention in detail. For those skilled in the art, without departing from the purpose and scope of the present invention and without unnecessary experiments, the present invention can be implemented within a relatively wide range under equivalent parameters, concentrations and conditions. Although specific embodiments of the present invention are given, it should be understood that the present invention can be further improved. In short, according to the principle of the present invention, this application intends to include any changes, uses or improvements to the present invention, including changes made using conventional techniques known in the art that are outside the scope disclosed in this application.

Claims

1. Use of any one of the following substances for regulating protein expression or regulating protein activity: P1. Use in regulating the grain length, grain width and / or 1000-grain weight of plant seeds; P2. Use in increasing the grain length, grain width and / or 1000-grain weight of plant seeds; P3. Use in regulating the spike length, number of spikelets and / or number of grains per spike of plant; P4. Use in increasing the spike length, number of spikelets and / or number of grains per spike of plant; P5. Use in regulating the yield per plant of plant; P6. Use in increasing the yield per plant of plant; P7. Use in regulating the protein content of plant seeds; P8. Use in increasing the protein content of plant seeds; P9. Use in high-yield breeding of plants and / or breeding of plants with high protein content in seeds; P10. Use in improving the quality of high-yield plants and / or plants with high protein content in seeds; The protein is as follows: A1) A protein with an amino acid sequence of SEQ ID NO: 9 in the sequence listing; A2) A protein with an amino acid sequence of SEQ ID NO: 3 in the sequence listing; A3) A protein with an amino acid sequence of SEQ ID NO: 6 in the sequence listing; A4) A protein obtained by substitution and / or deletion and / or addition of amino acid residues in the amino acid sequence shown in A1), A2) or A3), which is derived from A1), A2) or A3) and has the same function, or has more than 80% identity with the protein shown in A1), A2) or A3) and has the same function; A4) A fusion protein obtained by connecting a protein tag to the N-terminus and / or C-terminus of A1), A2) or A3).

2. The application according to claim 1, wherein: The protein is derived from wheat.

3. The application according to claim 1 or 2, characterized in that: The plant is any one of the following: D1) Dicotyledonous plants; D2) Monocotyledonous plants, D3) Plants of the order Poales, D4) Gramineous plants, D5) Plants of the genus Triticum, D6) Wheat.

4. Use of any one of the following biological materials related to the protein described in claim 1 or 2: Q1. Use in regulating the grain length, grain width and / or 1000-grain weight of plant seeds; Q2. Use in increasing the grain length, grain width and / or 1000-grain weight of plant seeds; Q3. Use in regulating the spike length, number of spikelets and / or number of grains per spike of plant; Q4. Use in increasing the spike length, number of spikelets and / or number of grains per spike of plant; Q5. Use in regulating the yield per plant of plant; Q6. Use in increasing the yield per plant of plant; Q7. Use in regulating the protein content of plant seeds; Q8. Use in increasing the protein content of plant seeds; Q9. Use in high-yield breeding of plants and / or breeding of plants with high protein content in seeds; Q10. Use in improving the quality of high-yield plants and / or plants with high protein content in seeds; The biological material is any one of the following B1) to B9): B1) A nucleic acid molecule encoding the protein described in 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); B5) A transgenic plant cell line containing the nucleic acid molecule described in B1), or a transgenic plant cell line containing the expression cassette described in B2); B6) A transgenic plant tissue containing the nucleic acid molecule described in B1), or a transgenic plant tissue containing the expression cassette described in B2); B7) A transgenic plant organ containing the nucleic acid molecule described in B1), or a transgenic plant organ containing the expression cassette described in B2); B8) A nucleic acid molecule that promotes or enhances the gene expression of the protein described in claim 1; B9) An expression cassette, recombinant vector, recombinant microorganism or transgenic plant cell line containing the nucleic acid molecule described in B8).

5. The application according to claim 4, wherein: The nucleic acid molecule described in B1) is the coding gene of the protein shown in any of the following b1), b2) or b3): b1) A cDNA molecule or DNA molecule whose coding sequence of the coding strand is the nucleotide of sequence 8, sequence 2 and / or sequence 5 in the sequence listing; b2) A DNA molecule whose nucleotide is sequence 7, sequence 1 and / or sequence 4 in the sequence listing, b3) A cDNA molecule or DNA molecule that hybridizes with the cDNA or DNA molecule defined in b2) and encodes a protein with the same function.

6. The application according to claim 4 or 5, characterized in that: The plant is any of the following: D1) Dicotyledonous plants; D2) Monocotyledonous plants, D3) Plants of the order Poales, D4) Gramineous plants, D5) Plants of the genus Triticum, D6) Wheat.

7. A method for increasing the grain length, grain width, 1000-grain weight, number of grains per spike, yield per plant and / or grain protein content of plants, including enhancing or increasing the activity of the protein described in claim 1 and / or the expression level of the coding gene of the protein described in claim 1 in the target plant, thereby increasing the grain length, grain width, 1000-grain weight, number of grains per spike, yield per plant and / or grain protein content of the target plant.

8. The method according to claim 7, characterized in that: The enhancement or increase of the activity of the protein described in claim 1 and / or the expression level of the coding gene of the protein described in claim 1 in the target plant is achieved by introducing the coding gene of the protein described in claim 1 into the target plant.

9. The method according to claim 7 or 8, characterized in that: The plant and / or the target plant is any of the following: D1) Dicotyledonous plants; D2) Monocotyledonous plants, D3) Plants of the order Poales, D4) Gramineous plants, D5) Plants of the genus Triticum, D6) Wheat.

10. The protein described in claim 1 and / or the biological material described in claim 4 or 5.

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