Application of rice vitamin B1 synthesis gene OsTHI1 or its coded protein in regulating drought resistance and / or regulating growth and development of rice

Abstract

The present application relates to a rice vitamin B1 synthesis gene OsTHI1 Or its encoded protein is applied to regulating drought resistance of rice and / or regulating growth and development of rice, and belongs to the technical field of genetic engineering. The rice vitamin B1 synthesis gene OsTHI1 The encoded protein sequence is shown as SEQ ID NO. 2. The present application researches and finds that OsTHI1 The positive regulation of drought resistance of rice: under drought stress, the overexpression plant shows stronger tolerance than the wild type, including higher survival rate, water retention capacity, chlorophyll content, and lower malondialdehyde content and up-regulation of multiple drought response genes. In addition, OsTHI1 It also regulates growth and development, maintains chloroplast integrity and photosynthesis. In summary, OsTHI1 The gene enhances the drought resistance of rice and regulates the growth and development, and provides a new gene resource for molecular breeding.

Description

Technical Field

[0001] This invention belongs to the field of genetic engineering technology, specifically relating to the rice vitamin B1 synthesis gene. OsTHI1 Or the application of its encoded proteins in regulating rice drought resistance and / or regulating rice growth and development. Background Technology

[0002] Vitamin B1 (thiamine, VB1), an indispensable component of the human diet, also plays a crucial role in plant growth and development; its deficiency can lead to seedling chlorosis and even death. Thiamine biosynthesis involves two branch pathways: the thiazole ring and the pyrimidine ring. Thiazole synthase THI1 is responsible for catalyzing the formation of the thiazole ring and is one of the key enzymes in thiamine synthesis. Rice (… Rice ) OsTHI1 Genes (also known as) OsDR8 ) is corn ( Zea mays Thiamine synthesis gene THI1 Homologous to. Existing research indicates that... OsTHI1 It mainly participates in the regulation of rice disease resistance and thiamine accumulation. Silencing is achieved using RNA interference technology. OsTHI1 Subsequently, the transgenic plants showed significantly reduced resistance to bacterial blight, a marked decrease in thiamine content, and suppressed expression levels of multiple defense response genes. Exogenous thiamine supplementation could partially restore the mutant's defense capabilities, indicating... OsTHI1 It plays an important role in the disease resistance and defense of rice. However, in addition to its disease resistance function, existing technologies have limited application in rice... THI1 Little is known about the functions of homologous genes in other biological processes. Summary of the Invention

[0003] The purpose of this invention is to address the shortcomings of existing technologies in rice cultivation. THI1 The current lack of homologous genes, aside from their disease resistance function, provides... THI1 A novel application of homologous genes in rice. Therefore, this invention provides a rice vitamin B1 synthesis gene. OsTHI1 Or the application of its encoded proteins in regulating rice drought resistance and / or regulating rice growth and development.

[0004] This invention provides a gene for synthesizing vitamin B1 in rice. OsTHI1 The application of its encoded protein in regulating rice drought resistance and / or regulating rice growth and development, said rice vitamin B1 synthesis gene OsTHI1 The amino acid sequence of the encoded protein is shown in SEQ ID NO.2.

[0005] Preferably, the application includes: overexpression of the rice vitamin B1 synthesis gene. OsTHI1 To improve the drought resistance of rice and / or improve the growth and development traits of rice.

[0006] This invention also provides an overexpression of the rice vitamin B1 synthesis gene. OsTHI1 The application of biomaterials in improving drought resistance in rice, including the rice vitamin B1 synthesis gene. OsTHI1 The amino acid sequence of the encoded protein is shown in SEQ ID NO.2; The biomaterial includes the rice vitamin B1 synthesis gene. OsTHI1 Recombinant expression vectors and / or containing the rice vitamin B1 synthesis gene OsTHI1 Recombinant microorganisms; The improvement of rice drought resistance includes any one or more of the following: ① to ⑥ ① Improve plant survival rate under drought stress; ② Improve the plant's recovery ability after rehydration under drought stress; ③ Reduce the water loss rate of detached leaves; ④ Increase leaf chlorophyll content under drought stress; ⑤ Reduce malondialdehyde content in leaves under drought stress; ⑥ Upregulate the expression levels of drought stress response genes; The drought stress response genes include OsRbcS1 , OsDip1 , OsLEA3 , OsERD1 , OsDREB2A and OsDREB2B At least one of them.

[0007] The present invention also provides a method for improving the drought resistance of rice, comprising the following steps: Overexpression of the rice vitamin B1 synthesis gene in rice OsTHI1 ; The rice vitamin B1 synthesis gene OsTHI1 The amino acid sequence of the encoded protein is shown in SEQ ID NO.2.

[0008] This invention also provides a method for cultivating highly drought-resistant rice, comprising the following steps: The transgenic rice obtained by the method described in claim 4 is used as a parent for breeding.

[0009] The present invention also provides a drought-resistant rice material in which the rice material overexpresses the rice vitamin B1 synthesis gene. OsTHI1 ; The rice vitamin B1 synthesis gene OsTHI1 The amino acid sequence of the encoded protein is shown in SEQ ID NO.2.

[0010] This invention also provides an overexpression of the rice vitamin B1 synthesis gene. OsTHI1The application of biomaterials in improving the growth and development traits of rice, including the rice vitamin B1 synthesis gene. OsTHI1 The amino acid sequence of the encoded protein is shown in SEQ ID NO. 2; The biomaterial includes the rice vitamin B1 synthesis gene. OsTHI1 Recombinant expression vectors and / or containing the rice vitamin B1 synthesis gene OsTHI1 Recombinant microorganisms; The improved rice growth and development traits include any one or more of the traits shown in ① to ④: ① Promotes chloroplast development; ②Enhance photosynthetic efficiency; ③ Increase grain length; ④ Increase starch content.

[0011] The present invention also provides a method for improving the growth and development traits of rice, comprising the following steps: Overexpression of the rice vitamin B1 synthesis gene in rice OsTHI1 ; The rice vitamin B1 synthesis gene OsTHI1 The amino acid sequence of the encoded protein is shown in SEQ ID NO.2; The improved rice growth and development traits include any one or more of the traits shown in ① to ④: ① Promotes chloroplast development; ②Enhance photosynthetic efficiency; ③ Increase grain length; ④ Increase starch content.

[0012] The present invention also provides a method for cultivating rice, comprising the following steps: The transgenic rice obtained by the method described in the above technical solution is used as a parent for breeding; The rice includes long-grain rice and / or high-starch rice.

[0013] The present invention also provides a rice material with improved agronomic traits, wherein the rice material overexpresses the rice vitamin B1 synthesis gene. OsTHI1 ; The rice vitamin B1 synthesis gene OsTHI1 The amino acid sequence of the encoded protein is shown in SEQ ID NO.2.

[0014] Beneficial effects: This invention provides a gene for synthesizing vitamin B1 in rice. OsTHI1 The application of its encoded protein in regulating rice drought resistance and / or regulating rice growth and development. The rice vitamin B1 synthesis gene described in this invention. OsTHI1The amino acid sequence of the encoded protein is shown in SEQ ID NO.2. This invention... OsTHI1 The gene promoter region contains three ABRE binding elements, and its expression level is regulated by the ABA signaling pathway. It can participate in the rice response to drought stress through the ABA signaling pathway.

[0015] This invention creates rice knockout mutants based on CRISPR-Cas9 technology. osthi1 And simultaneously by building OsTHI1 Overexpression vectors created overexpression materials OsTHI1-OE , discovered OsTHI1 The gene has the ability to positively regulate the resistance of rice to drought stress. Validated through examples, under drought stress conditions, OsTHI1 Overexpressing plants exhibited stronger tolerance than wild-type plants: after drought treatment and rehydration, the survival and recovery abilities of overexpressing plants were significantly better than those of wild-type plants; detached leaf water loss rate measurements showed that the water retention capacity of overexpressing plants was enhanced; under drought stress, the chlorophyll content of leaves from overexpressing plants was higher than that of wild-type plants, while the malondialdehyde (MDA) content was lower, indicating a milder degree of membrane lipid peroxidation damage; simultaneously, multiple drought stress response genes (such as...) were present in overexpressing plants. OsLEA3 , OsDREB2A The expression levels of (etc.) were significantly upregulated. These results confirm that... OsTHI1 Genes and their encoded proteins can enhance rice’s tolerance to drought stress, providing new gene resources and application directions for molecular breeding of drought-resistant rice.

[0016] Furthermore, the present invention also discovered OsTHI1 It participates in regulating the growth and development of rice. Transcriptome analysis and transmission electron microscopy observations show that... OsTHI1 Gene knockout mutants exhibit impaired chloroplast and thylakoid structures, disrupted stroma lamellae, and significantly reduced starch grain accumulation. Simultaneously, multiple genes involved in the core photosynthetic pathway (such as photosystem II) are also affected. psbO , psbQ , psbY , psb27 Optical System I psaD , psaF , psaH , psaL , psaN , psaO and the photosynthetic electron transport chain petE , petF The expression was significantly downregulated. This was verified by examples. osthi1 Knockout mutant seedlings exhibited an albino leaf phenotype, with significantly lower chlorophyll content and photosynthetic rate compared to the wild type; while OsTHI1The overexpressing plants exhibited normal chloroplast and thylakoid structures, significantly larger starch grain area than the wild type, increased instantaneous starch content in leaves, and significantly increased grain length. These results indicate that... OsTHI1 Genes and their encoded proteins can maintain the integrity of chloroplast structure, ensure normal photosynthesis, and thus regulate the growth and development traits of rice, providing new genetic resources for rice molecular breeding and quality improvement. Attached Figure Description To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the accompanying drawings used in the embodiments will be briefly described below.

[0017] Figure 1 for OsTHI1 A schematic diagram of the gene structure; Figure 2 for OsTHI1 Phylogenetic analysis diagram of this gene and other genes involved in thiamine biosynthesis; Figure 3 for OsTHI1 Figure 1 shows the construction and identification results of CRISPR-Cas9 and overexpression lines; where A represents the genome sequencing and identification results of the osthi1 knockout mutant line, and WT (wild type) serves as the control. osthi1#2 and osthi1#4 They are respectively osthi1 Strains 2 and 4; B is OsTHI1 Genes in osthi1 The image shows the expression level detection results in the knockout mutant lines. In the figure, L1~L11 represent osthi1 lines 1~11; C represents... OsTHI1 Schematic diagram of gene overexpression vector; D represents OsTHI1 Genes in OsTHI1-OE The graph shows the expression level detection results in the overexpression lines, where L1~L10 represent... OsTHI1-OE Strain 1~Strain 10; Figure 4 for OsTHI1 Gene expression pattern diagram; where A represents OsTHI1 A diagram of spatiotemporal gene expression patterns, where Root represents the root; Steam represents the stem; Leaf represents the leaf; S8 represents the pollen mother cell stage; S9 represents the meiotic stage; S10 represents the uninucleate stage; S11 represents the binucleate stage; S12 represents the pollen maturation stage; and B represents the OsTHI1 pro::GUS staining observation diagram. Figure 5 for OsTHI1 Phenotypic diagram of mutant plants (n=3), as shown in the figure represent PValue < 0.0001, WT indicates wild-type rice. osthi1-1 and osthi1-2 for osthi1 Knockout mutant lines OsTHI1-OVER-1 and OsTHI1-OVER-2 for OsTHI1 Overexpression lines; where A is OsTHI1 Phenotypic diagram of mutant plants, Bar = 15cm; B is OsTHI1 Leaf phenotypic diagram of the mutant, Bar=5cm; C is OsTHI1 Figure 1 shows the results of measuring the maximum photosynthetic rate of the mutant leaves; D represents... OsTHI1 Figure showing the results of chlorophyll content measurement in mutant leaves; Figure 6 This is a statistical chart of seed length and width (n>3). represent P Value < 0.01, represent P Value < 0.001; where A is the grain length phenotype (Bar = 1 cm); B is the grain width phenotype (Bar = 1 cm); C is the grain length statistical result chart; D is the grain width statistical result chart; E is the aboveground height statistical result chart 7 days after germination; F is the root length statistical result chart 7 days after germination. Figure 7 These are transmission electron micrographs (TEM) images of leaves. In the images, C represents chloroplasts, pg represents plastid globules, sg represents starch grains, st represents stroma, and th represents thylakoids. A through D are TEM images of wild-type leaves, with A and C having a Bar=2μm and B and D having a Bar=500nm. E (Bar=2μm) and F (Bar=500nm) are TEM images of osthi1-1 leaves; G (Bar=2μm) and H (Bar=500nm) are TEM images of osthi1-2 leaves; I (Bar=2μm) and J (Bar=500nm) are... OsTHI1-OVER-1 Transmission electron microscopy image of the leaf; K (Bar=2μm) and L (Bar=500nm) are... OsTHI1-OVER-2 Transmission electron microscope image of the blade; Figure 8 This is a graph showing the number and area of ​​starch granules (n>3). represent P Value < 0.01, represent P Value < 0.0001; where A is the result of starch granule quantity determination; B is the result of starch granule area determination; Figure 9 The figure shows the expression levels of genes related to vitamin B1 synthesis (n=3). represent P Value < 0.05, represent P Value < 0.01, represent P Value < 0.001, represent P Value < 0.0001; Figure 10 Image of leaf starch iodide staining (Bar=1 cm); Figure 11 This is a graph showing the expression of starch synthesis genes (n=3). represent P Value < 0.05, represent P Value < 0.01, represent P Value < 0.001, represent P Value < 0.0001; Figure 12 Tissue-specific expression staining of OsTHI1 pro::GUS before and after ABA treatment: a, leaf, Bar=1mm; b, magnified leaf, Bar=0.3mm; c, stem, Bar=1mm. Figure 13 for OsTHI1-OE Phenotypic images of rice plants under drought (Bar=10cm); where A is the phenotypic image of rice plants under drought treatment for 14 days; B is the phenotypic image of rice plants after 21 days of drought treatment followed by 7 days of rehydration. Figure 14 This is a graph showing the water loss rate of detached leaves (n=3); where, represent P Value < 0.05, represent P Value < 0.01; Figure 15 The graph shows chlorophyll content under drought stress (n=3); among them, represent P Value < 0.0001; Figure 16 The graph shows the malondialdehyde (MDA) content under drought stress (n=3); among which, represent P Value < 0.001, represent P Value < 0.0001; Figure 17 This is a graph showing gene expression in response to drought stress (n=3); among them, represent P Value < 0.05, represent P Value < 0.01, represent P Value < 0.001, represent P Value < 0.0001; Figure 18 For WT and osthi1 Statistical charts of expression level differences; where A is a bar chart of expression level differences; and B is a volcano plot of expression level differences. Figure 19 For WT and osthi1 GO classification analysis diagram of differentially expressed genes; Figure 20 For WT and osthi1 GO functional enrichment analysis diagram of differentially expressed genes; Figure 21 For WT and osthi1 KEGG classification analysis diagram of differentially expressed genes; Figure 22 For WT and osthi1 KEGG functional enrichment analysis diagram of differentially expressed genes; Figure 23 For WT and osthi1 KEGG functional enrichment analysis of differentially expressed genes in the photosynthetic energy metabolism pathway; where yellow background represents known genes / transcripts; green background represents new genes / transcripts; yellow + green represents known + new genes / transcripts; red borders represent upregulated genes; blue borders represent downregulated genes; and red + blue borders represent genes that are both upregulated and downregulated. Detailed Implementation

[0018] This invention provides a gene for synthesizing vitamin B1 in rice. OsTHI1 The application of its encoded protein in regulating rice drought resistance and / or regulating rice growth and development, said rice vitamin B1 synthesis gene OsTHI1 The amino acid sequence of the encoded protein is shown in SEQ ID NO.2.

[0019] As one embodiment, the rice vitamin B1 synthesis gene of the present invention OsTHI1 A schematic diagram of the gene structure is shown below. Figure 1 As shown. As one embodiment, the present invention... OsTHI1 The gene promoter region contains three ABRE-binding elements, and its expression level is regulated by the ABA signaling pathway, allowing it to participate in the rice's response to drought stress through this pathway. As one implementation method, this invention constructs a phylogenetic tree using the maximum likelihood method with the protein sequences encoded by SEQ ID NO.2 and other known thiamine biosynthesis genes, demonstrating that rice... OsTHI1 The gene is corn THI1 The genes are homologous, with a homology of 98%. As one implementation method, the phylogenetic tree described in this invention is as follows: Figure 2 As shown.

[0020] SEQ ID of the present invention NO.2 is specifically: MAAMATTASSLLKTSFAGARLPAAARNPTVSVAPRTGGAICNSISSSSSTPPYDLNAIRFSPIKESIVSREMTRRYMTDMITYADTDVVVVGAGSAGLSCAYELSKDPSVSVAVIEQSVSPGGGAWLGGQLFSAMVVRKPAHLFLDELGVAYDEQEDYVVIKHAALFTSTVMSR LLARPNVKLFNAVAVEDLIVKEGRVGGVVTNWALVSMNHDTQSCMDPNVMESRVVVSSCGHDGPFGATGVKRLQDIGMIDAVPGMRALDMNTAEDEIVRLTREVVPGMIVTGMEVAEIDGAPRMGPTFGAMMISGQKAAHLALKALGRPNAIDGTIKKAAAAAAHPELILASKDDGEIVDA.

[0021] As one embodiment, the rice vitamin B1 synthesis gene of the present invention OsTHI1

[0022] As one implementation method, the application of the present invention includes: overexpression of the rice vitamin B1 synthesis gene. OsTHI1 This invention aims to improve drought resistance and / or enhance growth and development traits in rice. As one implementation method, the invention constructs... OsTHI1 Overexpression vectors created overexpression materials OsTHI1-OE , discovered OsTHI1 The gene has the ability to positively regulate the ability of rice to resist drought stress. As one implementation method, the rice vitamin B1 synthesis gene described in this invention... OsTHI1 The overexpression material participates in regulating the growth and development of rice. OsTHI1-OE It can exert positive regulatory functions such as promoting chloroplast development, enhancing photosynthetic efficiency, increasing grain length, and increasing starch content.

[0023] This invention also provides an overexpression of the rice vitamin B1 synthesis gene. OsTHI1 The application of biomaterials in improving drought resistance in rice, including the rice vitamin B1 synthesis gene. OsTHI1 The amino acid sequence of the encoded protein is shown in SEQ ID NO.2; The biomaterial includes the rice vitamin B1 synthesis gene. OsTHI1 Recombinant expression vectors and / or containing the rice vitamin B1 synthesis gene OsTHI1 Recombinant microorganisms; The improvement of rice drought resistance includes any one or more of the following: ① to ⑥ ① Improve plant survival rate under drought stress; ② Improve the plant's recovery ability after rehydration under drought stress; ③ Reduce the water loss rate of detached leaves; ④ Increase leaf chlorophyll content under drought stress; ⑤ Reduce malondialdehyde content in leaves under drought stress; ⑥ Upregulate the expression levels of drought stress response genes; The drought stress response genes include OsRbcS1 , OsDip1 , OsLEA3 , OsERD1 , OsDREB2A and OsDREB2B At least one of them.

[0024] As one implementation method, the base vector of the recombinant expression vector of the present invention is a plant expression vector. p CAMBIA1300. As one embodiment, the recombinant expression vector of this invention uses Kpn I and Xbal I as double restriction sites, and the inserted fragment is... OsTHI1The open reading frame of the gene is used to construct a recombinant overexpression vector. As one embodiment, the recombinant microorganism of this invention includes a rice vitamin B1 synthesis gene. OsTHI1 The engineered bacteria. As one embodiment, the recombinant microorganism of the present invention contains the rice vitamin B1 synthesis gene. OsTHI1 The recombinant Agrobacterium. As one embodiment, the initial strain of the recombinant Agrobacterium of the present invention is Agrobacterium tumefaciens (…). Agrobacterium tumefaciens GV3101. As one implementation method, under drought stress conditions, the overexpression material described in this invention... OsTHI1-OE Compared to the wild type, it exhibited stronger tolerance: after drought treatment and rehydration, the survival and recovery ability of the overexpressing plants were significantly better than those of the wild type; the water loss rate of detached leaves showed that the water retention capacity of the leaves of the overexpressing plants was enhanced; under drought stress, the chlorophyll content of the leaves of the overexpressing plants was higher than that of the wild type, and the malondialdehyde (MDA) content was lower than that of the wild type, indicating that the degree of membrane lipid peroxidation damage was milder; at the same time, the expression levels of multiple drought stress response genes in the overexpressing plants were significantly upregulated.

[0025] The present invention also provides a method for improving the drought resistance of rice, comprising the following steps: Overexpression of the rice vitamin B1 synthesis gene in rice OsTHI1 ; The rice vitamin B1 synthesis gene OsTHI1 The amino acid sequence of the encoded protein is shown in SEQ ID NO.2.

[0026] As one embodiment, the present invention describes the overexpression of the rice vitamin B1 synthesis gene in rice. OsTHI1 By introducing the gene into rice OsTHI1 The overexpression vector is used to achieve this. As one implementation method, the present invention realizes the rice vitamin B1 synthesis gene through Agrobacterium-mediated genetic transformation. OsTHI1 Overexpression. As one implementation method, the base vector for the overexpression vector of the present invention is a plant expression vector. p CAMBIA1300. As one embodiment, the overexpression vector of this invention uses Kpn I and Xbal I as double restriction sites, and the inserted fragment is... OsTHI1 Open reading frames of genes were used to construct recombinant overexpression vectors.

[0027] This invention also provides a method for cultivating highly drought-resistant rice, comprising the following steps: The transgenic rice obtained by the method described in claim 4 is used as a parent for breeding.

[0028] The present invention also provides a drought-resistant rice material in which the rice material overexpresses the rice vitamin B1 synthesis gene. OsTHI1 ; The rice vitamin B1 synthesis gene OsTHI1 The amino acid sequence of the encoded protein is shown in SEQ ID NO.2.

[0029] As one embodiment, the rice material of the present invention includes rice plants, seeds, tissues, and callus tissue.

[0030] This invention also provides an overexpression of the rice vitamin B1 synthesis gene. OsTHI1 The application of biomaterials in improving the growth and development traits of rice, including the rice vitamin B1 synthesis gene. OsTHI1 The amino acid sequence of the encoded protein is shown in SEQ ID NO. 2; The biomaterial includes the rice vitamin B1 synthesis gene. OsTHI1 Recombinant expression vectors and / or containing the rice vitamin B1 synthesis gene OsTHI1 Recombinant microorganisms; The improved rice growth and development traits include any one or more of the traits shown in ① to ④: ① Promotes chloroplast development; ②Enhance photosynthetic efficiency; ③ Increase grain length; ④ Increase starch content.

[0031] As one embodiment, the recombinant microorganism of the present invention includes a rice vitamin B1 synthesis gene. OsTHI1 Engineered bacteria. As one implementation method, overexpression of the strains described in this invention... OsTHI1 Genes can maintain the integrity of rice chloroplast structure, ensure normal photosynthesis, and thus regulate the growth and development traits of rice. As one implementation method, transcriptomic analysis and transmission electron microscopy observations show that the genes described in this invention... OsTHI1 In gene knockout mutants, chloroplast and thylakoid structures are damaged, matrix lamellae are disrupted, and starch grain accumulation is significantly reduced. Simultaneously, multiple genes in the core photosynthetic pathway (such as photosystem II) are also affected. psbO , psbQ , psbY , psb27 Optical System I psaD , psaF , psaH , psaL , psaN , psaO and the photosynthetic electron transport chain petE , petF The expression is significantly downregulated. As one implementation method, the present invention... OsTHI1 Seedlings of the gene knockout mutant exhibited albino leaves, with significantly lower chlorophyll content and photosynthetic rate than the wild type; while OsTHI1 The overexpressing plants had normal chloroplast and thylakoid structures, significantly larger starch grain area than the wild type, increased instantaneous starch content in leaves, and significantly increased grain length.

[0032] The present invention also provides a method for improving the growth and development traits of rice, comprising the following steps: Overexpression of the rice vitamin B1 synthesis gene in rice OsTHI1 ; The rice vitamin B1 synthesis gene OsTHI1 The amino acid sequence of the encoded protein is shown in SEQ ID NO.2; The improved rice growth and development traits include any one or more of the traits shown in ① to ④: ① Promotes chloroplast development; ②Enhance photosynthetic efficiency; ③ Increase grain length; ④ Increase starch content.

[0033] As one embodiment, the present invention describes the overexpression of the rice vitamin B1 synthesis gene in rice. OsTHI1 By introducing the gene into rice OsTHI1 The overexpression vector is used to achieve this. As one implementation method, the present invention realizes the rice vitamin B1 synthesis gene through Agrobacterium-mediated genetic transformation. OsTHI1 Overexpression. As one implementation method, the base vector for the overexpression vector of the present invention is a plant expression vector. p CAMBIA1300. As one embodiment, the overexpression vector of this invention uses Kpn I and Xbal I as double restriction sites, and the inserted fragment is... OsTHI1 Open reading frames of genes were used to construct recombinant overexpression vectors.

[0034] The present invention also provides a method for cultivating rice, comprising the following steps: The transgenic rice obtained by the method described in the above technical solution is used as a parent for breeding; The rice includes long-grain rice and / or high-starch rice.

[0035] The present invention also provides a rice material with improved agronomic traits, wherein the rice material overexpresses the rice vitamin B1 synthesis gene. OsTHI1 ; The rice vitamin B1 synthesis gene OsTHI1 The amino acid sequence of the encoded protein is shown in SEQ ID NO.2.

[0036] As one embodiment, the rice material of the present invention includes rice plants, seeds, tissues, and callus tissue.

[0037] To further illustrate the present invention, the rice vitamin B1 synthesis gene provided by the present invention will be described below with reference to the accompanying drawings and embodiments. OsTHI1 The invention describes in detail the application of its encoded proteins in regulating drought resistance and / or regulating rice growth and development, but these descriptions should not be construed as limiting the scope of protection of this invention.

[0038] Example 1 osthi1 Obtaining and molecularly identifying knockout mutant plants Using the CRISPR-Cas9 method for osthi1 Mutants were generated, and the two target sequences used for mutagenesis were TGTCTCGCGAGATGACCCGGCGG (SEQ ID NO.3) and CCTGCGCGTACGAGCTCTCCAAG (SEQ ID NO.4). The recombinant expression vector described in this invention is a CRISPR-Cas9 gene editing vector, and the recombinant expression vector contains targeted... OsTHI1 The sgRNA coding sequence and PAM sequences of the gene were CGG and CCT, respectively. For each transformant (Agrobacterium-mediated transformation), a 200-300 bp region containing the target site was amplified at generation T0 for Sanger sequencing to screen for mutations. Sequencing confirmed homozygous lines. The mutants were created by Wuhan Aidijing Biotechnology Co., Ltd., and their verification was confirmed by sequencing. Real-time quantitative PCR was used to analyze the mutant plants. OsTHI1 Gene expression levels were detected. Real-time quantitative PCR was performed using the QIAGEN QuantiNova™ SYBR® Green PCR Kit on a QuantStudio 6 Flex real-time quantitative PCR platform. The rice actin gene OsActin (LOC_Os03g50885) was used as an internal control. Data analysis was conducted using a 2... -△△Ct Calculation by method. Select homozygous lines with lower expression. osthi1#2 and osthi1#4 ( Figure 3 Subsequent experiments were conducted on A and B in the sample, and the results were named as follows: osthi1-1 and osthi1-2 .

[0039] OsTHI1 Quantitative primers: OsTHI1 -F: CTGCAGGACATCGGCATGAT (SEQ ID NO.5); OsTHI1 -R: AGATCATCATGGCTCCGAACG (SEQ ID NO. 6).

[0040] Example 2 OsTHI1 Obtaining and molecularly identifying overexpressing plants 1. OsTHI1 Construction of overexpression vectors Using rice Nipponbare cDNA as a template, upstream primer 5'-GAATTCGAGCTCGGTACCATGGCAGCCATGGCCAC-3' (SEQ ID NO.7) and downstream primer 5'-GCCTGCAGGTCGACTCTAGATCACTTGTCATCGTCATCCTTGTAATCG-3' (SEQ ID NO.8) were used to amplify the cDNA via RT-PCR. OsTHI1 The open reading frame (ORF) was determined. After separation of the PCR products by agarose gel electrophoresis, the target DNA fragment was recovered. The pCAMBIA1300 vector was digested with Kpn I and Xbal I restriction enzymes, and the digestion products were recovered. The recovered fragment was then compared with... p CAMBIA 1300 vector ligation to obtain recombinant vectors p CAMBIA 1300- OsTHI1 (like Figure 3 (C in the text). Completed by Nanjing Genscript Biotech Co., Ltd.

[0041] 2. OsTHI1 Preparation of overexpression plants Using Nipponbare rice as background material, and employing materials containing... p CAMBIA 1300- OsTHI1 The Agrobacterium GV3101 vector was used to infect rice to obtain T0 generation transgenic plants overexpressing the gene (Wuhan Aidijing Biotechnology Co., Ltd.). DNA was extracted from the T0 generation rice using the TPS method. The selective marker gene HPT (hygromycin phosphotransferase gene) on the vector was amplified by PCR to detect positive plants. The primer sequences were HPT-F: 5'-AGAAGAAGATGTTGGCGACCT-3' (SEQ ID NO.49) and HPT-R: 5'-GTCCTGCGGGTAAATAGCTG-3' (SEQ ID NO.50). Positive plants were self-pollinated to the T2 generation, and homozygous transgenic lines were obtained through hygromycin resistance selection. The T3 generation plants were used for real-time quantitative PCR detection. OsTHI1 Gene expression level (detection method same as in Example 2, primers are) OsTHI1-F / OsTHI1-R ), the result is as follows Figure 3 As shown in D. Two T3 generation lines with high expression levels were selected. OsTHI1-OE#1 and OsTHI1-OE#2 Named OsTHI1-OE-1 and OsTHI1-OE-2 , for use in subsequent experiments.

[0042] Example 3 OsTHI1 Obtaining and verifying pro::GUS plants Will OsTHI1 The promoter region of the gene (2000 bp upstream of the start codon ATG) was cloned into the GUSplus vector pBI101.3 GUSplus using SalI / KpnI (this process was commissioned to Nanjing Genscript Biotech Co., Ltd.). After obtaining the sequencing report, the accuracy of the vector was verified by enzyme digestion. Rice was transformed by Agrobacterium infection to obtain GUS transgenic plants (Wuhan Aidijing Biotechnology Co., Ltd.). Rice DNA was extracted using the TPS method. The NPT II gene on the GUS vector was used as a selection marker. PCR positive detection was performed using primers Npt II_F: 5'-CGGTGCCCTGAATGAACTCCAG-3' (SEQ ID NO.51) and Npt II_R: 5'-GCCATGTGTCACGACGAGATCCTC-3' (SEQ ID NO.52). The size of the bands in the electrophoretic amplification products determined the T0 generation positive plants. The T0 generation positive plants were continuously self-pollinated and identified by NPT II resistance screening to obtain the T3 generation homozygous lines for subsequent GUS histochemical staining analysis.

[0043] Example 4 OsTHI1 Gene expression pattern analysis Real-time quantitative PCR experiments were performed on the roots, stems, leaves, and flowers at different developmental stages of wild-type rice to detect... OsTHI1 Gene expression levels. The results showed... OsTHI1 It is expressed most highly in stems and leaves, and is expressed at all stages of flower development, especially in the mid-microspore stage (S11). OsTHI1 The gene expression level is the highest ( Figure 4 (A in the middle).

[0044] right OsTHI1 pro::GUS GUS histochemical staining was performed on different tissues of positive plants to observe the promoter expression patterns. Results are as follows: Figure 4 As shown in Figure B, the Bar value for germinating seedlings is 5 mm, the Bar value for the entire anther is 1 mm, the Bar value for a single anther is 1 mm, the Bar value for the glumes is 2 mm, and the Bar value for leaves is 500 μm. GUS staining signals were successfully detected in the embryo and bud at 3, 7, and 10 days after rice germination. Furthermore, strong GUS signals were observed in the embryo, bud, anther, glumes, and leaves, but almost no expression was observed in the roots.

[0045] Example 5 OsTHI1 Mutant phenotypic analysis 1. OsTHI1 Mutant phenotypic analysis Select plump rice seeds (Nipponbare wild type and) OsTHI1 Transgenic lines were dehulled and placed in wide-mouthed bottles filled with water. They were then cultured in a water-jacketed constant-temperature incubator in the dark at 35℃ for 2 days until the seeds showed white sprouts, with the water changed twice daily. The sprouted seeds were then evenly sown on a homemade mesh tray, placed in rice nutrient solution, and cultured in a light incubator with the following parameters: 28℃ / light level 6 / 12 h, 22℃ / light level 0 / 12 h. At the 4-leaf stage, the seeds were transplanted into soil and planted outdoors in Kunming, Yunnan Province, China. During winter, they were grown in a greenhouse at an average temperature of 22℃~28℃. The growth of different plants at various stages of development was observed and compared. Figure 5 (A and B in the original text). Discovery osthi1 Compared to the wild type, the mutant exhibits whitishness in the fourth leaf at the four-leaf stage, followed by gradual whitening of the other leaves. As the number of whitish leaves increases, all mutant plants eventually die, while the overexpression material displays a phenotype similar to the wild type.

[0046] When the mutant rice reached the four-leaf stage, a 10 mM vitamin B1 solution was sprayed on the leaves every two days. When the mutant rice reached the tillering stage, a 50 mM vitamin B1 solution was sprayed on the leaves every two days. It was found that after exogenous supplementation with vitamin B1 solution, the chlorotic phenotype of the mutant was restored, the white leaves gradually turned green, and the plant could grow normally.

[0047] To verify whether vitamin B1 affects chlorophyll synthesis, this study measured the chlorophyll content and photosynthetic rate of leaves from wild-type and mutant plants. Chlorophyll content was determined using the ethanol immersion method, and photosynthetic rate was measured using a chlorophyll fluorometer. The results showed that... osthi1 The content of chlorophyll a, chlorophyll b, and carotenoids in the leaves was significantly lower than that in the wild type, and the photosynthetic rate was also significantly lower than that in the wild type. osthi1 It was 83% lower than the wild type. This indicates that... osthi1 Impaired chlorophyll synthesis ( Figure 5 (C and D in the text).

[0048] To understand whether the seed length and width of genetically modified plants were affected, samples were collected. osthi1 and OsTHI1-OE Statistical observation of seeds ( Figure 6 (A and B in the original text), and the results showed that, OsTHI1-OE The grain length of both overexpression lines was significantly higher than that of the wild type. Figure 6 (C in the text), but the particle width of knockout mutants and overexpression mutants was not significantly different from that of wild type (C). Figure 6(D in the text). The aboveground height and underground height were measured 10 days after rice germination. Figure 6 The E and F values ​​(in the wild type) showed no difference compared to the wild type. These results indicate... OsTHI1 Gene overexpression can lead to increased grain length in rice.

[0049] 2. Observation using transmission electron microscopy To investigate whether the chloroplast structure of the mutant was affected, transmission electron microscopy (TEM) was used to observe the chloroplasts and thylakoid structures of the leaves. Figure 7 TEM results show osthi1 The chloroplasts and thylakoids are damaged, the stroma lamellae are disrupted, there is almost no starch grain accumulation, and there are few plastid globules. OsTHI1-OE Both overexpression lines exhibited phenotypes similar to the wild type, and OsTHI1-OE The average area of ​​starch granules in the two strains was significantly higher than that in the wild type. Figure 8 The results showed that the vitamin B1 synthesis gene... OsTHI1 Mutations damage the chloroplast structure of plants, leading to abnormal chloroplast function and ultimately causing plant death.

[0050] Example 6 Assay of Vitamin B1 Synthesis-Related Gene Expression Levels To compare the changes in the expression levels of genes related to vitamin B1 synthesis with those in the wild type, the expression of genes involved in thiamine synthesis in the mutant was determined using real-time quantitative PCR. Figure 9 Primer sequences are shown in Table 1.

[0051] Table 1 Primers for Real-Time Quantitative PCR

[0052] Research found that osthi1 In mutants, OsTHI1 Downregulation of gene expression, vitamin B1 synthesis gene OsTH1 , OsPALE1 and OsTPK1 All were significantly revised upwards. OsTHIC and OsTPK3 The expression of these genes was significantly downregulated in the plants, while the expression of these genes in overexpressed plants showed the opposite trend. The results indicate that... OsTHI1 Gene mutations can lead to changes in the thiamine synthesis gene. OsTHIC、OsTH1 , OsPALE1 , OsTPK1 and OsTPK3 Significant changes occur in gene expression.

[0053] In Arabidopsis thaliana, AtTHI1It can interact with CPK33 and inhibit CPK33 kinase activity. This study determined... OsCPK12 exist osthi1 Expression in mutants. The results showed that... osthi1 In knockout mutants, OsCPK12 Gene expression was significantly upregulated, while OsTHI1-OE In mutants, OsCPK12 Gene expression was significantly downregulated. This indicates... OsTHI1 Genes have an effect OsCPK12 Gene expression.

[0054] Example 7 OsTHI1 Genes influence starch synthesis right osthi1 and OsTHI1-OE Observation of mutant leaves by iodine staining ( Figure 10 The results showed that at the end of the day, wild-type and OsTHI1-OE Iodine staining of leaves produces a deep blue color, while osthi1 The leaves are light brown; at the end of darkness, the wild type and OsTHI1-OE They all appear dark blue, while osthi1 The leaves are light blue, and the color is lighter than that of the wild type. These results indicate that... OsTHI1 It is crucial for transient starch synthesis in rice leaves. OsTHI1 Genes are key genes in starch synthesis.

[0055] To understand the expression of starch synthesis genes, the expression of starch synthesis genes in rice leaves was studied. OsLESV、OsESV1、 AGPL1、AGPL2 and AGPL3 Real-time quantitative PCR detection was performed. Figure 11 The primer sequences are shown in Table 1. Experimental results showed that... OsTHI1-OE Among the plants, OsLESV , OsESV1 The expression of [a specific substance] was significantly downregulated. These results indicate that... OsTHI1 Genes influence the expression of starch synthesis genes.

[0056] Example 8 OsTHI1 Response to drought stress analysis 1. OsTHI1 The expression of is regulated by the ABA signaling pathway. Among known downstream transcription factors, members of the bZIP family, specifically the ARB (ABA response element binding protein) / ABF (ABRE binding factor) subfamily, play a crucial role in ABA signaling. The cistropic regulatory elements of gene promoters were analyzed using an online analysis website (https: / / bioinformatics.psb.ugent.be / webtools / plantcare / html / ), and the results are shown in Table 2.

[0057] Table 2 OsTHI1 Gene promoter analysis

[0058] Discover OsTHI1 Three ABRE-binding elements were found (Table 2). This experiment used 50 μmol ABA for 12 hours of hydroponic treatment. OsTHI1 After pro::GUS plants were established, GUS staining was performed on leaves and stems. The experimental results showed ( Figure 12 Under normal conditions, this gene shows weak expression in leaves and stems; however, after ABA treatment, the GUS staining signal was significantly enhanced, suggesting that... OsTHI1 The expression level of is regulated by the ABA signaling pathway.

[0059] 2. OsTHI1-OE Rice plants enhance drought stress tolerance ABA regulatory elements (such as ABRE) are present in the promoter regions of many drought-responsive genes. Under drought stress, increased ABA levels activate genes containing ABA regulatory elements, including some genes associated with drought tolerance. The proteins encoded by these genes may participate in various physiological processes to enhance plant tolerance to drought stress.

[0060] wild type, OsTHI1-OE Rice plants and OsTHIC-OE Rice plants ( OsTHIC The gene locus LOC_Os03g47610, belonging to the same genus as rice thiamine synthesis pathway genes, was used as a control material in the same experiment. It was planted in grid-shaped soil pots, germinated, and grew for 50 days. After 14 days of drought (without watering), OsTHI1-OE The seedlings of both strains exhibited less curling than the wild type and... OsTHIC-OE plant ( Figure 13 ), and when the soil was re-irrigated (watered to soil saturation capacity) for 7 days after a 21-day drought, it was found that OsTHI1-OE The plants are growing better than the wild type and OsTHIC-OE The plant. The above results indicate that... OsTHI1 Overexpression can significantly enhance the tolerance of rice to drought stress.

[0061] 3. Determination of water loss rate of detached leaves Plants in water-scarce environments regulate transpiration from their leaves to slow water loss. To compare WT and... OsTHI1-OE To determine whether there were significant differences in the water loss rate of detached leaves of rice plants, this study measured... OsTHI1-OE Water loss per hour within the first 4 hours after detachment of rice leaves. Results showed ( Figure 14 ), at 2 h, OsTHI1-OE The water loss rates of both overexpression lines were significantly lower than those of the wild type, but at 4 h, there was no significant difference between their water loss rates and those of the wild type. These results indicate that... OsTHI1-OE The plant has improved the water retention capacity of its leaves to some extent, reducing water loss.

[0062] 4. Chlorophyll content determination under drought stress Under drought stress, plants face challenges such as stomatal closure and reduced carbon dioxide supply. Higher chlorophyll content helps rice absorb as much light energy as possible under limited water conditions, maintaining a certain level of photosynthesis. Even when drought causes a decrease in stomatal conductance and intercellular carbon dioxide concentration, chlorophyll ensures sufficient ATP and NADPH production in the light-dependent reactions, providing energy and reducing power for subsequent Calvin cycle reactions, thus maintaining a certain photosynthetic carbon assimilation efficiency and ensuring the plant can synthesize enough organic matter for its own growth and to cope with stress. Chlorophyll content was determined using the ethanol immersion method. This study found (… Figure 15 Under drought stress, OsTHI1-OE The leaves of overexpressing plants contained higher levels of chlorophyll a, chlorophyll b, and carotenoids than the wild type.

[0063] 5. Determination of malondialdehyde (MDA) content under drought stress Under drought stress, lipid peroxidation of plant cell membranes intensifies, leading to oxidative damage. Malondialdehyde (MDA), as one of the main end products of lipid peroxidation, can quantify the degree of cell damage induced by drought stress to some extent, thus serving as a key indicator for assessing the damage caused by drought stress to plants. From the perspective of plant stress physiology, the dynamic changes in MDA content reflect the effectiveness of the plant's cellular antioxidant defense system and its ability to maintain cell membrane stability during drought stress, indirectly reflecting the strength of the plant's adaptation and resistance to drought stress. MDA content was determined using the thiobarbituric acid method. This study found that ( Figure 16 Under drought stress, OsTHI1-OE The malondialdehyde (MDA) content in the overexpressing plants was significantly lower than that in the wild type. OsTHI1- OE The overexpression resulted in less severe membrane lipid oxidation damage in the plants, indicating that... OsTHI1 It helps reduce oxidative damage to plants under high osmotic stress.

[0064] 6. OsTHI1-OE Analysis of drought stress response gene expression in plants In order to understand OsTHI1-OE This experiment investigated the expression levels of drought-resistance-related genes in overexpressed plants using real-time quantitative PCR. OsRbcS1 , OsDip1 , OsLEA3 , OsERD1 , OsDERB2A and OsDERB2B exist OsTHI1-OE The expression level in overexpressed rice plants ( Figure 17 (The primer sequences are shown in Table 1. It was found that the expression levels of most drought genes were significantly higher than those of wild type.)

[0065] Example 9 OsTHI1 Transcriptome analysis 1. Differentially expressed gene analysis Take wild type and osthi1 The aboveground parts of knockout mutant rice seedlings at the four-leaf stage were sequenced by Shanghai Meiji Biotechnology Co., Ltd. Total RNA extraction was performed using a Qiagen kit. Sequencing experiments were conducted using the Illumina Stranded mRNA Prep, Ligation (constant) method for library construction, followed by sequencing on the NovaSeq X Plus platform. Statistical results of differentially expressed genes are shown below. Figure 18 As shown in the figure, DEGs represent differentially expressed genes. The expression difference of DEGs is at least two-fold, with a p-adjustment value < 0.05. Genes were screened using an adjusted p-value (p-adjust) < 0.05 and a fold change > 2 as thresholds to obtain statistical analysis of expression differences and volcano plots. A total of [number missing] genes were screened. osthi1 There were 210 upregulated genes, 203 downregulated genes, and 38,580 indifferential genes.

[0066] 2. GO classification and enrichment analysis of differentially expressed genes In order to investigate osthi1 To determine which biological processes are affected, we performed a Gene Ontology (GO) classification analysis on differentially expressed genes (DEGs). The results are as follows: Figure 19As shown, in terms of biological processes, differentially expressed genes are mainly concentrated in metabolism, cellular responses, stimuli, and biological regulation. At the cellular component level, related differentially expressed genes are mainly enriched in cellular structural entities and protein complexes. In terms of molecular function, differentially expressed genes are mainly concentrated in catalytic activity and binding function.

[0067] To further clarify the specific processes involved in differentially expressed genes, GO enrichment analysis was performed in this experiment. The results are as follows: Figure 20 As shown, 27 differentially expressed genes were enriched in photosynthesis, 22 in the photosystem, 49 in chloroplasts, and 53 in plastids. During carbohydrate metabolism, 14 genes were upregulated and 15 were downregulated. We also detected 11 differentially expressed genes enriched in the abscisic acid and jasmonic acid signaling pathways, mainly including… RSOsPR10 , OsPYL / RCAR4 , RPR10b , OsPR10a The expression of these genes was downregulated. These results indicate that in osthi1 Among the differentially expressed genes produced by gene knockout mutants, some genes are directly or indirectly involved in photosynthesis in chloroplasts and plant metabolism.

[0068] 3. KEGG classification and enrichment analysis of differentially expressed genes In order to investigate OsTHI1 This study investigates the differential gene expression and functions induced by gene knockout in plants. Using KEGG (Kyoto Encyclopedia of Genes and Genomes) analysis, it delves into the metabolic pathways involved by differentially expressed genes, aiming to reveal their crucial roles in plant metabolism. KEGG taxonomic enrichment analysis results are presented below. Figure 21 The data shows that 158 ​​differentially expressed genes were annotated in the metabolism category, including 37 in energy metabolism, 28 in carbohydrate metabolism, 27 in amino acid metabolism, and 18 in cofactor and vitamin metabolism. In the genetic information processing category, there were 6 differentially expressed genes for signal transduction and 3 for membrane transport. In the environmental information processing category, there were 3 differentially expressed genes for translation, 2 for folding, sorting and degradation, and 1 for chromosome.

[0069] based on Figure 22 Functional annotation analysis of the KEGG pathway database showed that differentially expressed genes were significantly enriched in multiple photosynthesis-related metabolic pathways. Twelve differentially expressed genes are involved in core photosynthetic processes, of which eight genes are specifically located in the PSII antenna protein complex pathway, and another eleven genes are significantly enriched in the photosynthetic carbon assimilation pathway.

[0070] 4. OsTHI1 Participating in photosynthesis Combining the albino phenotype of the mutant with sequencing analysis of transcriptome data, it was found that OsTHI1 It is associated with energy metabolism, carbohydrate metabolism, and amino acid metabolism. Therefore, this study selected the photosynthetic energy metabolism pathway for KEGG pathway analysis of differentially expressed genes. Log2 fold change in transcriptome data is used to describe changes in gene expression levels and is an important indicator for measuring gene expression changes; positive values ​​indicate upregulation, and negative values ​​indicate downregulation. For example... Figure 23 As shown in Table 3, which describes the differentially expressed genes and regulatory directions in the photosynthetic pathway, no genes were upregulated. Four genes in PSII were downregulated: PsbO (LOC_Os01g31690) PsbQ (LOC_Os07g36080) PsbY (LOC_Os08g02630) and Psb27 (LOC_Os03g21560). Six genes were downregulated in PSⅠ, namely... PsaD (LOC_Os08g44680) PsaF ( OsPS1 -F, LOC_Os03g56670), PsaH (LOC_Os05g48630) PsaL (LOC_Os12g23200) PsaN (LOC_Os12g08770) and PsaO (LOC_Os04g33830). Two genes are downregulated in light and electron transport, namely... PetE ( OsCPL1 (LOC_Os06g01210) and PetF ( OsFd1 (LOC_Os08g01380). In summary, OsTHI1 Gene knockout mutations alter the metabolic pathways of rice, and OsTHI1 It participates in photosynthesis, thereby affecting starch synthesis in rice.

[0071] Table 3. Differentially expressed genes and regulatory directions in the photosynthetic pathway.

[0072] Although the above embodiments have provided a detailed description of the present invention, they are only some embodiments of the present invention, and not all embodiments. People can obtain other embodiments based on these embodiments without creative effort, and these embodiments all fall within the protection scope of the present invention.

Claims

1. Rice vitamin B1 synthesis gene OsTHI1 The application of its encoded protein in regulating rice drought resistance and / or regulating rice growth and development, said rice vitamin B1 synthesis gene OsTHI1 The amino acid sequence of the encoded protein is shown in SEQ ID NO.

2.

2. The application according to claim 1, characterized in that, The applications include: overexpression of the rice vitamin B1 synthesis gene. OsTHI1 To improve the drought resistance of rice and / or improve the growth and development traits of rice.

3. Overexpression of the rice vitamin B1 synthesis gene OsTHI1 The application of biomaterials in improving drought resistance in rice, including the rice vitamin B1 synthesis gene. OsTHI1 The amino acid sequence of the encoded protein is shown in SEQ ID NO.2; The biomaterial includes the rice vitamin B1 synthesis gene. OsTHI1 Recombinant expression vectors and / or containing the rice vitamin B1 synthesis gene OsTHI1 Recombinant microorganisms; The improvement of rice drought resistance includes any one or more of the following: ① to ⑥ ① Improve plant survival rate under drought stress; ② Improve the plant's recovery ability after rehydration under drought stress; ③ Reduce the water loss rate of detached leaves; ④ Increase leaf chlorophyll content under drought stress; ⑤ Reduce malondialdehyde content in leaves under drought stress; ⑥ Upregulate the expression levels of drought stress response genes; The drought stress response genes include OsRbcS1 , OsDip1 , OsLEA3 , OsERD1 , OsDREB2A and OsDREB2B At least one of them.

4. A method for improving the drought resistance of rice, characterized in that, Includes the following steps: Overexpression of the rice vitamin B1 synthesis gene in rice OsTHI1 ; The rice vitamin B1 synthesis gene OsTHI1 The amino acid sequence of the encoded protein is shown in SEQ ID NO.

2.

5. A method for cultivating highly drought-resistant rice, characterized in that, Includes the following steps: The transgenic rice obtained by the method described in claim 4 is used as a parent for breeding.

6. A drought-resistant rice material, characterized in that, The rice material overexpressed the rice vitamin B1 synthesis gene. OsTHI1 ; The rice vitamin B1 synthesis gene OsTHI1 The amino acid sequence of the encoded protein is shown in SEQ ID NO.

2.

7. Overexpression of the rice vitamin B1 synthesis gene OsTHI1 The application of biomaterials in improving the growth and development traits of rice, including the rice vitamin B1 synthesis gene. OsTHI1 The amino acid sequence of the encoded protein is shown in SEQ ID NO.2; The biomaterial includes the rice vitamin B1 synthesis gene. OsTHI1 Recombinant expression vectors and / or containing the rice vitamin B1 synthesis gene OsTHI1 Recombinant microorganisms; The improved rice growth and development traits include any one or more of the traits shown in ① to ④: ① Promotes chloroplast development; ②Enhance photosynthetic efficiency; ③ Increase grain length; ④ Increase starch content.

8. A method for improving the growth and development traits of rice, characterized in that, Includes the following steps: Overexpression of the rice vitamin B1 synthesis gene in rice OsTHI1 ; The rice vitamin B1 synthesis gene OsTHI1 The amino acid sequence of the encoded protein is shown in SEQ ID NO.2; The improved rice growth and development traits include any one or more of the traits shown in ① to ④: ① Promotes chloroplast development; ②Enhance photosynthetic efficiency; ③ Increase grain length; ④ Increase starch content.

9. A method for cultivating rice, characterized in that, Includes the following steps: Breeding is carried out using the transgenic rice obtained by the method described in claim 8 as a parent; The rice includes long-grain rice and / or high-starch rice.

10. A rice material with improved agronomic traits, characterized in that, The rice material overexpressed the rice vitamin B1 synthesis gene. OsTHI1 ; The rice vitamin B1 synthesis gene OsTHI1 The amino acid sequence of the encoded protein is shown in SEQ ID NO.2.

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