Application of OsGTF5 gene in regulating rice plant brittleness, plant height and heading date

Abstract

The application discloses application of an OsGTF5 gene in regulating rice plant brittleness, plant height and / or heading stage, and belongs to the technical field of rice breeding. The coding sequence of the OsGTF5 gene is shown as SEQ ID No. 1. In specific application, the OsGTF5 gene of wild-type rice is silenced through a mutation or knockout mode, and the obtained rice strain has low mechanical strength, low plant height and / or delayed heading stage; or the OsGTF5 gene is transformed into the rice strain with the silenced OsGTF5 gene, and the obtained genetically complementary rice strain has restored mechanical strength, plant height and / or heading stage. The gtf5 mutant is identified through a Mutmap method, and it is proved through experimental research that the OsGTF5 gene is a gene for regulating rice plant brittleness, height and heading stage. The application provides valuable resources for breeding of new rice brittleness varieties and exploration of a heading stage regulation mechanism, and has important significance in utilization of renewable biomass resources and the like.

Description

TECHNICAL FIELD

[0001] The application relates to application of an OsGTF5 gene in regulating rice plant brittleness, plant height and heading stage, and belongs to the technical field of rice breeding. BACKGROUND

[0002] Rice is widely planted in East Asia, South Asia, the Middle East, the West Indies and Latin America, and there are more than 110,000 local varieties, providing more than one-fifth of the global human calorie consumption. As an important food crop, the yield-related agronomic traits of rice, such as heading stage, stem strength and efficient use of post-harvest straw, have always been of concern.

[0003] Rice straw is an important renewable biomass resource with a wide range of applications, such as fermentation for feed, biomass fuel, soil for cultivation and raw materials for various substances such as bio-oil. At present, the main straw treatment methods are burning, enzymatic hydrolysis and microbial degradation, which pollute the environment and have low utilization efficiency. Efficient use of rice straw is an important problem currently faced in rice production.

[0004] The heading stage is a key trait for the transformation of vegetative growth into reproductive growth, and is closely related to the photoperiod sensitivity and thermosensitivity of rice. At present, the main breakthrough point for unrestricted planting of rice in different regions and different seasons is to breed varieties with long basic vegetative growth period and weak photoperiod sensitivity, which is an important goal of rice breeding. It is closely related to the yield of rice.

[0005] Plants have evolved tough cell walls to resist physical and chemical damage in nature, which is the main reason for the low recovery rate of straw. The strength of plant stems depends on the composition and structural properties of the cell wall. The main components of the plant cell wall are cellulose, hemicellulose, lignin and pectin. The cellulose molecules provide tensile strength, the intermolecular hydrogen bonds between adjacent cellulose molecules make them strongly adhere to each other in overlapping parallel arrangement, forming highly ordered crystal aggregates (cellulose microfibrils), these crystal aggregates are further bonded by hemicellulose, pectin regulates ion balance, adheres cells and maintains cell wall integrity signaling, lignin and pectin fill the gaps to form a complex network, further enhancing its tensile strength and rigidity. The structure of mature plant cell wall is divided into primary wall, secondary wall and middle lamella, among which the secondary wall is the main determinant of mechanical support force, therefore the composition of the secondary wall of the brittle mutant may have changed, leading to its inability to maintain the integrity of the cell wall function. The brittle rice mutant plant has the characteristics of reduced mechanical strength and easy crushing of straw, and is an excellent new material for breeding rice varieties. At the same time, the delay of the heading stage of rice increases the length of the basic vegetative growth, which lays a rich material foundation for the reproductive growth in the later period.

[0006] So far, the reported rice brittle mutants usually show that the loss of genes interferes with the normal biosynthesis of the secondary wall, which changes the content of cellulose and lignin in the cell wall, resulting in the decrease of the mechanical strength of rice stem, such as BC15, BC19 genes are related to the synthesis of cellulose; the loss of BC14 gene leads to the change of fiber bundle arrangement, etc. These studies lay the foundation for exploring the regulation network of rice plant brittleness, but the specific mechanism and its use in breeding need to be studied. It is of great significance to explore the genes that are insensitive to light and temperature and regulate the heading date of rice.

[0007] In summary, the rice brittle mutant provides a new perspective for the research of straw efficient utilization, and a new direction for the research of heading date regulation, which has important significance for the selection of new varieties and the development and utilization of germplasm resources in the future. SUMMARY

[0008] In view of the above prior art, the application provides the application of OsGTF5 gene (Xylan arabinosyltransferase) in regulating the brittleness, plant height and / or heading date of rice plant.

[0009] The application is realized by the following technical solutions:

[0010] The application of OsGTF5 gene in regulating the brittleness, plant height and / or heading date of rice plant, the coding sequence of the OsGTF5 gene is shown as SEQ ID No. 1, and the amino acid sequence of the OsGTF5 protein is shown as SEQ ID No. 2.

[0011] Further, in specific application, the OsGTF5 gene of wild type rice is silenced or the expression amount of the OsGTF5 gene is reduced by mutation or knockout, and the obtained rice strain has lower mechanical strength, shorter plant height and / or delayed heading date compared with the wild type;

[0012] Or: the coding region or the complete OsGTF5 gene of the OsGTF5 gene is transformed in the rice strain with silenced OsGTF5 gene, or the expression amount of the OsGTF5 gene is increased, and the obtained genetic complementary rice strain has restored mechanical strength, plant height and / or heading date time compared with the rice strain with silenced OsGTF5 gene.

[0013] Further, the mutation is selected from EMS mutagenesis; and the knockout is selected from CRISPR-Cas9 gene editing technology knockout.

[0014] Further, the nucleotide sequence of the coding region of the mutated OsGTF5 gene is shown in SEQ ID No. 3, in which the A at position 136 is mutated to T; or shown in SEQ ID No. 5, in which two bases at positions 90 and 91 are deleted; or shown in SEQ ID No. 6, in which three bases at positions 53, 90 and 91 are deleted.

[0015] Further, the nucleotide sequence of the complete OsGTF5 gene is shown in SEQ ID No. 4, which comprises the complete 5'UTR, exon, intron and 3'UTR (from 2793 bp before ATG to 1225 bp after TAG).

[0016] Further, the rice variety is Nipponbare (Nip).

[0017] The application of the OsGTF5 protein in regulating the plant brittleness, plant height and / or heading time of rice, wherein the amino acid sequence of the OsGTF5 protein is shown in SEQ ID No. 2.

[0018] Further, in a specific application, the OsGTF5 gene of the wild-type rice is silenced or the expression amount of the OsGTF5 gene is reduced by mutation or knockout, and the obtained rice strain has lower mechanical strength, shorter plant height and / or delayed heading time compared with the wild-type.

[0019] Or: the coding region of the OsGTF5 gene or the complete OsGTF5 gene is transformed into the OsGTF5 gene silenced rice strain, or the expression amount of the OsGTF5 gene is increased, and the obtained genetically complemented rice strain has restored mechanical strength, plant height and / or heading time compared with the OsGTF5 gene silenced rice strain.

[0020] The mutated OsGTF5 gene, wherein the nucleotide sequence of the coding region is shown in SEQ ID No. 3, or shown in SEQ ID No. 5, or shown in SEQ ID No. 6.

[0021] The application of the mutated OsGTF5 gene or related biological material in preparing transgenic rice. The transgenic rice has lower mechanical strength, shorter plant height and / or delayed heading time. The related biological material includes a recombinant vector containing the mutated OsGTF5 gene and a transformant containing the mutated OsGTF5 gene.

[0022] A method for regulating the fragility of rice plants, the height of rice plants and / or the heading stage: silencing or reducing the expression of the OsGTF5 gene of wild-type rice by mutation or knockout, and obtaining a rice strain, compared with the wild type, the mechanical strength of the rice plant is lower, the plant height is shorter and / or the heading stage is delayed.

[0023] Or: transforming the coding region of the OsGTF5 gene or the complete OsGTF5 gene in the OsGTF5 gene silenced rice strain, or increasing the expression of the OsGTF5 gene, and obtaining a genetically complementary rice strain, compared with the OsGTF5 gene silenced rice strain, the mechanical strength of the rice plant, the plant height and / or the heading stage time are restored.

[0024] Further, the mutation is selected from EMS mutagenesis; the knockout is selected from CRISPR-Cas9 gene editing technology knockout.

[0025] The OsGTF5 gene has NCBI accession number LOC4342012, and its coding sequence is shown as SEQ ID No. 1. The OsGTF5 gene contains 2 exons and 1 intron, and only one transcript, encodes 460 amino acids, and the amino acid sequence is shown as SEQ ID No. 2, and the encoded protein is located in the Golgi body in the cytoplasm. After conservative domain prediction and analysis, the OsGTF5 protein is likely to be a xylan arabinose transferase, which contains a hidden Markov model PF04577, and belongs to the GT61 family (Glycosyltransferase family 61, GT61) of glycosyltransferase.

[0026] The present application identifies a gtf5 mutant by Mutmap technology. In the mutant, the 136th A in the coding region of the OsGTF5 gene (on the second exon) is mutated to T, causing premature termination of translation, and the corresponding coding sequence is shown as SEQ ID No. 3. The mutant plant shows the phenotypes of shorter plant height, lower mechanical strength, easy breaking of stems and leaves, and delayed heading stage. In agricultural production, the straw of the mutant is more easily crushed than the wild type, which has important significance for improving the utilization rate of renewable biomass, and has great application value in biomass conversion, pulp and paper industry; in breeding, the mutant has a longer vegetative growth period, which provides new materials for future breeding of new varieties.

[0027] The application constructs a complementary vector pCAMBIA1300-OsGTF5 for the OsGTF5 gene, and genetically transforms a gtf5 mutant to obtain positive lines Com1 and Com2, and the plant height, leaf and stem strength, and heading period of the positive lines are restored. The application further constructs a knockout vector for the gene mutation through a CRISPR-Cas9 system, and genetically transforms a wild type Nipponbare to obtain positive lines KO1 and KO2, and it is observed that the positive lines exhibit similar phenotypes to the mutant, i.e., the plant height is short, the stem and leaf are easy to break, and the heading period is delayed. These experiments further prove that the loss of the OsGTF5 gene is the main reason for the brittleness, short plant height, and delayed heading period of the gtf5 mutant plant, and the OsGTF5 gene is a gene for regulating the brittleness and heading period of a rice plant, and can be used for preparing a transgenic plant. The application has important significance for crop improvement and improvement of the resistance to lodging, and has a wide application prospect in future agricultural product processing. The application provides valuable resources for the cultivation of a new rice brittleness variety and the exploration of a heading period regulation mechanism, and has important significance in the utilization of renewable biomass resources. BRIEF DESCRIPTION OF DRAWINGS

[0028] Figure 1 : Comparison diagram of plant height and internode length of wild type (Nip) and gtf5 mutant, wherein A: plant at the jointing stage, scale = 10 cm; B: plant at the filling stage, scale = 10 cm; C: plant at the heading stage, scale = 10 cm; D: comparison of internode length, scale = 1 cm, 5 cm.

[0029] Figure 2 : Comparison diagram of plant height of wild type (Nip), gtf5 mutant, complementary lines (Com1, Com2), and OsGTF5 gene knockout lines (KO1, KO2), wherein A: plants of wild type, gtf5 mutant, and complementary lines, scale = 10 cm; B: plants of wild type, gtf5 mutant, and OsGTF5 gene knockout lines, scale = 10 cm.

[0030] Figure 3 : Diagram of sequencing verification result of the complementary lines, and the mutation base position exhibits a double peak.

[0031] Figure 4 : Diagram of sequencing verification result of the gene knockout lines, and the mutation position exhibits a base deletion.

[0032] Figure 5 : Diagram of subcellular localization of the OsGTF5 protein, wherein A is an OsGTF5-GFP fusion protein, B is a Golgi Marker, and C is a combined image of A and B.

[0033] Figure 6Figure 6: Comparison of plant height among wild type (Nip), gtf5 mutant, complemented lines (Coml, Com2) and OsGTF5 gene knockout lines (KO1, KO2). * indicates P<0.05, ** indicates P<0.01, *** indicates P<0.001.

[0034] Figure 7 Figure 7: Comparison of internode length among wild type (Nip), gtf5 mutant, complemented lines and OsGTF5 gene knockout lines. * indicates P<0.05, ** indicates P<0.01, *** indicates P<0.001; A: first internode length; B: second internode length; C: third internode length; D: fourth internode length.

[0035] Figure 8 Figure 8: Comparison of leaf and stem strength among wild type (Nip), gtf5 mutant, complemented lines and OsGTF5 gene knockout lines. * indicates P<0.05, ** indicates P<0.01, *** indicates P<0.001; A: first internode strength assay; B: second internode strength assay; C: third internode strength assay; D: fourth internode strength assay; E: first leaf strength assay; F: second leaf strength assay; G: wild type (Nip), gtf5 mutant, complemented lines and OsGTF5 gene knockout lines first leaf strength phenotype; H: wild type (Nip), gtf5 mutant, complemented lines and OsGTF5 gene knockout lines third internode strength phenotype.

[0036] Figure 9 Figure 9: Comparison of heading date among wild type (Nip), gtf5 mutant, complemented lines (Coml, Com2) and OsGTF5 gene knockout lines (KO1, KO2). * indicates P<0.05, ** indicates P<0.01, *** indicates P<0.001. DETAILED DESCRIPTION

[0037] The application will be further described with reference to the following examples. However, the scope of the present application is not limited to the following examples. Those skilled in the art will understand that various changes and modifications can be made to the present application without departing from the spirit and scope of the present application.

[0038] The instruments, reagents, materials involved in the following examples are all conventional instruments, reagents, materials available in the prior art, and can be obtained through regular commercial channels, unless otherwise specified. The experimental methods involved in the following examples are generally carried out according to conventional experimental conditions or according to the experimental conditions recommended by the manufacturer, unless otherwise specified.

[0039] Example 1: Obtaining rice gtf5 mutant

[0040] The rice gtf5 mutant is from a EMS mutagenesis library of japonica rice Nipponbare, and is stable after 5 generations of continuous selfing. The mutant shows plant brittleness (mechanical strength is low, stems and leaves are easy to break), dwarfing and delayed heading. The comparison of plant height and internode length between the rice gtf5 mutant and wild-type Nipponbare is shown in Table 1. Figure 1 As shown in Table 1, the plant height and internode length of the rice gtf5 mutant at the jointing stage, filling stage and heading stage are lower than those of wild-type Nipponbare. Figure 1

[0041] Further genetic analysis of the gtf5 mutant was performed using a hybrid combination prepared from japonica rice Nipponbare. The F1 plant phenotype did not separate, and normal plant height and heading stage plants were separated from the F2 population, and plants with brittleness, dwarfing and delayed heading were classified into two categories, 155 and 46, respectively. Chi-square test showed a 3:1 segregation ratio (χ 2 = 0.48), indicating that the brittleness, dwarfing and delayed heading of the gtf5 mutant plant are controlled by a pair of recessive nuclear genes.

[0042] Example 2: Mutmap method for identifying gtf5 mutant

[0043] To identify the control gene causing plant brittleness, dwarfing and delayed heading, the gtf5 mutant was backcrossed with the wild type, and the Mutmap method was used to identify the plants showing brittleness in the backcross F2 separation population. According to the candidate gene, referring to the LOC_Os06g49320 locus information of the rice whole genome database (http: / / rice.uga.edu / cgi-bin / sequence_display.cgi?orf=LOC_Os06g49320.1), a pair of amplification primers OsGTF5-seq-F and OsGTF5-seq-R (see Table 1) were designed for PCR amplification, and the complete coding region sequence containing OsGTF5 was obtained and compared with the wild type sequence.

[0044] The sequence difference between the mutant and Nipponbare was obtained by resequencing, an InDel marker was designed for accurate positioning, and the physical interval was shortened. Finally, the coding region (second exon) of LOC_Os06g49320 gene was sequenced, and the 136th A was mutated to T, resulting in premature termination of translation. The coding sequence of the mutant is shown in SEQ m No. 3.

[0045] Table 1 Primer sequences used in the experiment

[0046] ​

[0047] Example 3: OsGTF5 gene function identification

[0048] To confirm that the phenotype of gtf5 mutant is caused by the loss of function of OsGTF5 gene, genetic complementation and OsGTF5 gene knockout experiments were performed on gtf5 mutant and wild type, respectively. Details are as follows.

[0049] (1) The full-length sequence of OsGTF5 gene was obtained by PCR amplification using Nipponbare genomic DNA as template and primers OsGTF5-EcoRI-F and OsGTF5-EcoRI-R (see Table 1), followed by homologous recombination with EcoRI-digested pCAMBIA1305 vector, transformation of E. coli, and sequencing verification of the plaque. The plasmid with correct sequence was used to transform Agrobacterium tumefaciens EHA105 for transgenic complementation verification of gtf5 mutant. The plant genetic transformation was completed by Wuhan Boyuan Biotechnology Co., Ltd. Finally, the positive plants of genetic complementation were obtained, which showed similar phenotype to wild type, i.e. the plant height, stem and leaf strength, and heading period were restored. The comparison of plant height is shown in Figure 2 A.

[0050] (2) CRISPR-Cas9 system was used to construct OsGTF5 gene knockout strain. The vector for creating knockout strain by CRISPR-Cas9 was pCAMBIA1300-SK-5G, which was purified and recovered by AarI digestion. A target site was designed on the first exon of rice OsGTF5 gene to knockout OsGTF5 gene. The sequence of sgRNA designed according to the target sequence was 5'-GATGTAGGTGAGGACGCAGA-3', and the Oligo DNA primers designed for the target site were OsGTF5-AarI-F and OsGTF5-AarI-R (see Table 1). The primers were reacted at 100°C for 5 min and cooled at room temperature. The vector and fragment were ligated using T4 ligase, transformed into E. coli, and the plaque was sequenced. The plasmid with correct sequence was used to transform Agrobacterium tumefaciens EHA105, and the plant genetic transformation was completed by Wuhan Boyuan Biotechnology Co., Ltd. Nipponbare was used as the transformation receptor, and finally the positive plants of OsGTF5 gene editing were obtained, which showed similar phenotype to gtf5 mutant, i.e. the plant height became shorter, the stem and leaf strength decreased, and the heading period delayed. The comparison of plant height is shown in Figure 2 B.

[0051] The complementation strain and OsGTF5 gene knockout strain constructed above were subjected to sequencing verification, and the results are shown in Figure 3 、 Figure 4As shown, the mutation base position of the complementary line shows a double peak, and the mutation position of the OsGTF5 gene knockout line shows 2 base deletions and 3 base deletions.

[0052] Example 4: Subcellular localization of OsGTF5 protein

[0053] According to the NCBI conserved domain prediction (https: / / www.ncbi.nlm.nih.gov / Structure / cdd / wrpsb.cgi), OsGTF5 contains a DUF563 conserved domain, and the prediction by the LocTree 3 (https: / / www.rostlab.org / services / loctree3 / ) website shows that it is subcellularly localized to the Golgi apparatus. Subcellular localization experiments were performed, and a subcellular localization vector was constructed in this embodiment, and the vector construction process is as follows:

[0054] Using primers OsGTF5-GFP-EcoRI-F and OsGTF5-GFP-EcoRI-R (see Table 1), the full-length cDNA of the OsGTF5 gene (i.e., the sequence shown in SEQ ID NO. 1) was used as a template for amplification, and then the fragment was recombined into the EcoRI site in the pYBA1132 vector (the pYBA1132 vector was donated by Dr. Yao Lei of Beijing Academy of Agriculture and Forestry Sciences) by homologous recombination, and finally the OsGTF5-GFP recombinant vector was formed. In tobacco cells, transient expression was performed, and fluorescence signals were detected by confocal microscopy, and the experimental results showed that the OsGTF5-GFP fusion protein was localized to the Golgi apparatus, as shown in Figure 5 .

[0055] Example 5: Plant height determination of wild type, gtf5 mutant, complementary line and knockout line

[0056] Plant height is a manifestation of cell morphology. The wild type Nipponbare (31 strains), gtf5 mutant (28 strains), complementary line (Com1: 51 strains; Com2: 48 strains) and knockout line (KO1: 41 strains; KO2: 30 strains) were measured for plant height, and the plant height data was subjected to significant difference comparison analysis by T-test. * indicates t-test Pvalue < 0.05; ** indicates t-test Pvalue < 0.01; *** indicates t-test Pvalue < 0.001. The statistical results are as follows: Figure 6As shown in Table 2, the average plant heights of wild-type, gtf5 mutant, complementary lines (Com1, Com2), and knockout lines (KO1, KO2) were 79.7 cm, 72.6 cm, 80.8 cm, 82.7 cm, 70.0 cm, and 68.2 cm, respectively. Compared with wild-type, the plant heights of gtf5 mutant and knockout lines decreased by an average of 7.1 cm, 9.7 cm, and 11.5 cm, respectively, with significant differences, while the plant height of complementary lines did not change significantly.

[0057] Table 2. Statistical data of traits

[0058] Traits Nip gtf5 Com1 Com2 KO1 KO2 Plant height / cm 79.7±2.7 72.6±4.0 80.9±3.8 82.7±3.7 70.0±2.7 68.2±4.3 First internode length / cm 2.2±0.8 1.8±0.7 2.1±0.8 2.6±1.1 1.5±0.7 1.6±0.9 Second internode length / cm 7.1±1.3 6.0±1.3 7.2±1.3 7.7±1.5 4.5±1.4 5.0±1.1 Third internode length / cm 13.3±1.4 12.2±1.1 14.3±2.3 14.0±1.3 10.9±1.3 11.7±1.1 Fourth internode length / cm 32.2±1.6 28.6±2.2 33.8±3.8 33.8±1.6 30.3±2.2 30.4±2.2 First internode brittleness / N 78.2±27.4 15.9±4.7 57.39±27.1 63.5±34.4 12.0±5.4 17.5±8.1 Second internode brittleness / N 115.4±10.9 24.3±11.3 106.1±19.5 118.8±9.0 22.9±11.8 30.3±13.8 Third internode brittleness / N 110.5±11.7 25.9±10.7 94.5±20.4 104.2±19.0 27.0±15.2 29.3±11.3 Fourth internode brittleness / N 66.9±20.1 14.4±6.6 57.1±14.2 54.2±15.8 15.8±5.9 16.7±6.0 First leaf brittleness / N 13.2±3.4 3.1±1.5 14.3±3.5 14.8±3.1 ＜1 ＜1 Second leaf brittleness / N 12.3±2.9 1.6±0.6 11.6±3.5 12.4±2.6 ＜1 ＜1 Heading date / d 64.1±1.2 69.4±1.4 65.5±1.9 64.5±1.2 74.5±3.1 70.7±1.4

[0059] Statistics on the length of each intersegment are as follows: Figure 7 As shown in Table 2, from Figure 7 As shown in Table 2, compared with the wild type, the number of internodes in the gtf5 mutant remained unchanged, but the length of each internode was significantly shorter. Compared with the gtf5 mutant, the internode lengths of the complementary lines were restored. Specifically, the average lengths of the first internodes of Nip, gtf5, Com1, Com2, KO1, and KO2 were 2.2cm, 1.8cm, 2.1cm, 2.6cm, 1.5cm, and 1.6cm, respectively; the average lengths of the second internodes were 7.1cm, 6.0cm, 7.2cm, 7.7cm, 4.5cm, and 5.0cm, respectively; the average lengths of the third internodes were 13.3cm, 12.2cm, 14.3cm, 14.0cm, 10.9cm, and 11.7cm, respectively; and the average lengths of the fourth internodes were 32.2cm, 28.6cm, 33.8cm, 33.8cm, 30.3cm, and 30.4cm, respectively.

[0060] Example 6: Measurement of stem and leaf mechanical strength of wild-type, gtf5 mutant, complementary lines and knockout lines

[0061] Mechanical strength reflects the lodging resistance of plants. The mechanical strength of leaves and stems of wild-type Nipponbare, the gtf5 mutant, complementary lines, and knockout lines was measured, and the differences were compared using a T-test. Statistical results are shown below. Figure 8, the first internode mechanical strength of 27 strains of Nip, 21 strains of gtf5, 16 strains of Coml, 36 strains of Com2, 18 strains of KOl and 24 strains of KO2 were measured; the second internode mechanical strength of 43 strains of Nip, 32 strains of gtf5, 27 strains of Coml, 44 strains of Com2, 42 strains of KOl and 36 strains of KO2 were measured; the third internode mechanical strength of 40 strains of Nip, 33 strains of gtf5, 24 strains of Coml, 44 strains of Com2, 40 strains of KOl and 37 strains of KO2 were measured; the fourth internode mechanical strength of 40 strains of Nip, 33 strains of gtf5, 24 strains of Coml, 38 strains of Com2, 47 strains of KOl and 35 strains of KO2 were measured. The results showed that the mechanical strength of gtf5 mutants and knockout strains were significantly reduced compared with the wild type, and the mechanical strength of the complementary strains was significantly restored compared with the gtf5 mutants. Similarly, the mechanical strength of the first leaf of 37 strains of Nip, 38 strains of gtf5, 27 strains of Coml and 39 strains of Com2 were measured; the mechanical strength of the second leaf of 52 strains of Nip, 38 strains of gtf5, 27 strains of Coml and 39 strains of Com2 were measured, and the results showed that the gtf5 leaf mutants exhibited significantly reduced brittleness compared with Nip and complementary strains, and the leaf mechanical strength of knockout strains was less than 1 N, which was difficult to detect.

[0062] Example 7: Statistics of heading date of wild type, gtf5 mutants, complementary strains and knockout strains

[0063] The heading date of wild type Nipponbare (37 strains), gtf5 mutants (31 strains), complementary strains (23 strains of Coml; 23 strains of Com2) and knockout strains (30 strains of KOl; 30 strains of KO2) were counted (Fuyang, Hangzhou), and the heading date data were analyzed for significant difference by T-test. The statistical results are shown in Table 2. Figure 9 , the heading date of Nip, gtf5 mutants, Coml, Com2, KOl and KO2 were 64.1 days, 69.4 days, 65.5 days, 64.5 days, 74.5 days and 70.7 days, respectively, and the heading date of mutants and knockout strains was significantly delayed compared with the wild type, by 5.3 days, 10.4 days and 6.3 days, respectively.

[0064] The above examples are provided to those skilled in the art to fully disclose and describe how to implement and use the claimed embodiments, rather than to limit the scope of the disclosure herein. Modifications obvious to those skilled in the art will be within the scope of the appended claims.

Claims

1. Application of mutation or knockout of OsGTF5 gene in rice during delayed heading stage, wherein the coding sequence of OsGTF5 gene is shown in SEQ ID No.1, and the expression level of OsGTF5 gene is silenced or reduced by mutation or knockout of OsGTF5 gene in rice.

2. The application according to claim 1, characterized in that: The mutation was selected from EMS mutagenesis; the knockout was selected from CRISPR-Cas9 gene editing technology knockout.

3. The application according to claim 1, characterized in that: The nucleotide sequence of the coding region of the mutated OsGTF5 gene is shown in SEQ ID No. 3, 5 or 6.

4. The application according to any one of claims 1 to 3, characterized in that: The rice variety mentioned is Nipponbare.

5. A method for delaying the heading stage of rice, characterized in that: The OsGTF5 gene in wild-type rice is silenced by mutation or knockout, or the expression level of the OsGTF5 gene is reduced; the coding sequence of the OsGTF5 gene is shown in SEQ ID No. 1.

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