Genetic engineering application of rice ethylene response transcription factor OsKTR2

Abstract

The invention discloses genetic engineering application of a rice ethylene response transcription factor OsKTR2. The invention discloses a rice ethylene response transcription factor OsKTR2, and the CDS sequence of the rice ethylene response transcription factor OsKTR2 is SEQ ID NO.1. In the invention, it is found for the first time that a recombinant expression vector constructed by the OsKTR2 gene is transfected into wild rice (Oryza sativa. Ssp.cv.Japonica) through an agrobacterium-mediated rice transgenic technology, and the effective tiller number, the total tiller number and the maturing rate of rice are increased, so that the yield of rice is increased. In addition, the expression of the OsKTR2 is induced to be up-regulated by low potassium stress, so that the potassium absorption of the rice is promoted.

Description

Technical Field

[0001] The invention belongs to the technical field of genetic engineering, and relates to the genetic engineering application of rice ethylene response transcription factor OsKTR2. Background Art

[0002] Potassium (K) is one of the important mineral macronutrients and is widely involved in many physiological processes of plant growth and development, including osmotic regulation, enzyme activation, ion balance, stomatal movement, and resistance to biotic and abiotic stresses (Wang et al., 2013; Anschütz et al., 2014). As an important inorganic osmotic ion, K + It can regulate the osmotic potential of plant cells and play an important role in plant resistance to osmotic / drought stress. K can also regulate plant photosynthesis and the transfer and metabolism of carbohydrates, ultimately determining the yield and quality of crops. When K supply is insufficient, plants will show yellowing of leaves due to water loss, weak plants, and sensitivity to abiotic stresses such as drought, which seriously affects plant growth and development (Munson, 1985). On the contrary, when K supply is sufficient, plants can enhance their resistance to biotic and abiotic stresses.

[0003] It has been reported that low potassium stress can induce potassium deficiency signals in root epidermal cells, including Ca 2+ , ROS, plant hormones, microRNAs, etc. These signals are further transmitted and trigger the expression of downstream target genes (especially K channels and high-affinity K transporters) through transcription factors, ultimately achieving plant adaptation to low potassium environments (Wang and Wu, 2013).

[0004] AP2 / ERF is one of the largest plant transcription factor families, containing 119-200 family members (Du et al. 2014; Nakano et al. 2006; Rao et al. 2015; Zhuang et al. 2008), which are involved in physiological processes such as plant ethylene signal transduction, stress response, metabolism, and response to external stimuli (Han et al., 2016; Koyama et al. 2013; Lee et al. 2012; Li et al. 2007; Fits and Memelink 2000; Trujillo et al. 2008; Zhu et al. 2014). AP2 / ERF superfamily members all contain a conserved AP2 binding domain consisting of approximately 57-66 amino acids, which is composed of three β-folded strands (DNA binding domain) and one α-helical element (Okamuro et al., 1997; Danny WK Ng et al., 2018). According to the difference in the number of copies of the AP2 domain among the members, we usually divide AP2 / ERF transcription factors into four subfamilies: AP2, ERF, RAV and Soloist (Nakano et al. 2006; Licausi et al. 2010a). However, different members of this family often play corresponding biological functions in only one link of physiological processes such as plant ethylene signal transduction, stress response, metabolism and response to external stimuli. Kim et al. found that RAP2.11, a member of the Vb subfamily of Arabidopsis ERF transcription factors, improves the tolerance of plants to low potassium environments by regulating the expression of the high-affinity K transporter AtHAK5 gene and other related genes in the low potassium signal transduction pathway (Kim et al., 2012). The expression of other members of the AP2 / ERF family, such as AtERF98, MsERF8, JcERF011, and CaERFLP1, can improve the salt tolerance of plants (Chen et al. 2012; Lee et al. 2004; Tang et al. 2016; Zhang et al. 2004, 2012b). In addition, a small number of AP2 / ERF members can respond to two or more environmental stresses at the same time. It is unpredictable what biological function a new AP2 / ERF member has. Summary of the invention

[0005] The purpose of the present invention is to provide a genetic engineering application of rice ethylene response transcription factor OsKTR2, mainly that the self-promoter overexpression material can increase the tillering number, improve the fruit setting rate and yield.

[0006] The purpose of the present invention is achieved through the following technologies:

[0007] The cDNA sequence of rice ethylene-responsive transcription factor OsKTR2 is shown in SEQ ID NO.1.

[0008] A recombinant expression vector comprising the rice ethylene-responsive transcription factor OsKTR2 according to claim 1.

[0009] The recombinant expression vector is pCAMBIA1305 vector.

[0010] The recombinant expression vector is further obtained by inserting the rice ethylene response transcription factor OsKTR2 described in claim 1 into the SacI and PstI restriction sites of the starting vector pCAMBIA1305, then using EcoRI and SacI to remove the Ubiquitin promoter and inserting the OsKTR2 gene's own promoter.

[0011] The application of the rice ethylene response transcription factor OsKTR2 is selected from any one or more of the following:

[0012] (1) Increase the total number of tillers, effective tillers, and fruit setting rate of crops;

[0013] (2) Increase rice yield;

[0014] (3) Promote rice's absorption of potassium;

[0015] The use of the rice ethylene response transcription factor OsKTR2 in constructing transgenic rice, wherein the transgenic rice has any one or more of the following traits:

[0016] (1) Increase the total number of tillers, effective tillers, and fruit setting rate of crops;

[0017] (2) Increase rice yield;

[0018] (3) Promote rice's absorption of potassium;

[0019] The application of the recombinant expression vector is selected from any one or more of the following:

[0020] (1) Increase the total number of tillers, effective tillers, and fruit setting rate of crops;

[0021] (2) Increase rice yield;

[0022] (3) Promote rice's absorption of potassium;

[0023] The use of the recombinant expression vector in constructing transgenic rice, wherein the transgenic rice has any one or more of the following traits:

[0024] (1) Increase the total number of tillers, effective tillers, and fruit setting rate of crops;

[0025] (2) Increase rice yield;

[0026] (3) Promote rice's absorption of potassium;

[0027] Beneficial effects of the present invention:

[0028] 1. In the present invention, a recombinant expression vector constructed by the rice ethylene-responsive transcription factor OsKTR2 was discovered for the first time in the world, and was transfected into the wild-type rice Oryza sativa.ssp.cv. Japonica through Agrobacterium-mediated rice transgenic technology. We found that the total tiller number, effective tiller number, fruit setting rate and grain yield of the OsKTR2 transgenic rice material were significantly improved compared with those of the wild-type rice.

[0029] 2. The OsKTR2 gene in the present invention is derived from rice, and the promoter is also the sequence upstream of the OsKTR2 gene, which is not an exogenous gene, and therefore has biosafety. The constructed rice ethylene-responsive transcription factor OsKTR2 plant expression vector can be directly used for Agrobacterium-mediated plant genetic transformation to obtain new germplasm that can increase the total tillering and effective tillering of plants.

[0030] 3. The overexpression of the OsKTR2 gene in the present invention can promote the absorption of potassium by rice, especially under low potassium conditions. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] Figure 1 , Molecular identification and plant type screening of OsKTR2 transgenic rice materials

[0032] Figure A: Phenotypes of Nipponbare wild-type rice WT and OsKTR2 self-promoter overexpressing material Pro-KTR2; Figure B: The expression level of OsKTR2 in Pro-KTR2 relative to WT; Figures CD: Biomass of roots and aboveground parts of WT and Pro-KTR2 materials.

[0033] Figure 2 , subcellular localization of OsKTR2 and its expression in different growth stages and tissues of rice

[0034] Figure A: The expression of OsKTR2 in different growth stages and tissues of Nipponbare wild-type rice; Figure B: OsKTR2 is localized in the nucleus of rice cells.

[0035] Figure 3 OsKTR2 expression is upregulated by potassium deficiency, H2O2 and ACC stress

[0036] Figure AB: The expression levels of OsKTR2 gene in the root and aboveground parts at different time points after being subjected to 0.05mM K stress; Figure CD: The expression levels of OsKTR2 gene in the root and aboveground parts at different time points after being subjected to 10mM H2O2 stress; Figure EF: The expression levels of OsKTR2 gene in the root and aboveground parts at different time points after being subjected to 100μMACC stress.

[0037] Figure 4 Effects of OsKTR2 expression on potassium absorption and accumulation in rice

[0038] Figure AB: Phenotypes of WT and Pro-KTR2 materials after treatment with 0.05mM K and 1mM K, respectively; Figure CD: Biomass of WT and Pro-KTR2 materials after treatment with 0.05mM K and 1mM K, respectively; Figure EF: Phenotypes of WT and Pro-KTR2 materials after treatment with 0.05mM K and 1mM K, respectively + concentration; Figure GH: total K content of WT and Pro-KTR2 materials after treatment with 0.05 mM K and 1 mM K, respectively.

[0039] Figure 5 , OsKTR2 transgenic materials and wild-type rice field test phenotypes

[0040] Figure A: Agronomic traits, yield and tiller number of WT and Pro-KTR2 materials in low-potassium soil; Figure B: Agronomic traits, yield and tiller number of WT and Pro-KTR2 materials in normal potassium soil.

[0041] Figure 6 , map of pCAMBIA1305 expression vector.

[0042] Figure 7 , map of pCAMBIA1305.1-GFP expression vector. DETAILED DESCRIPTION

[0043] Example 1. Cloning of OsKTR2 gene

[0044] 1. Template: RNA was extracted from leaves of Nipponbare (NB) wild-type rice grown in normal hydroponics for 2 weeks and reverse transcribed into cDNA, which was used as a template for cloning the OsKTR2-CDS sequence.

[0045] 2. PCR primer design: Find the gene sequence of OsKTR2 on the NCBI website (https: / / www.ncbi.nlm.nih.gov / ), design primers using the primer design software Primer5.0, add SacI (GAGCTC) and PstI (CTGCAG) restriction site sequences to the 5' and 3' ends of the primers, and add a sequence on the pCAMBIA1305 plasmid vector to form a 41bp homologous recombination primer (F1 and R1).

[0046] Upstream primer F1:

[0047] 5'-ATTTACGAACGATAG GAGCTC ATGGAGCTCAACTTCCAAGT-3' (SEQ ID NO.2);

[0048] Downstream primer R1:

[0049] 5'-GTGATTTTTGCGGAC CTGCAG CTAATGGCACAGTGGCGAGA-3' (SEQ ID NO. 3).

[0050] 3. PCR amplification of OsKTR2 gene: PCR Buffer 2μl, dNTP Mix 2μl, upstream and downstream primers 1μl each, template 1μl, KOD high-fidelity enzyme 1μl, redistilled water 13μl. The PCR amplification program is as follows: 94℃ pre-denaturation for 5min, 94℃ denaturation for 30s, 62℃ annealing for 30s, 72℃ renaturation extension for 1min, after 35 cycles, 72℃ 10min full extension,

[0051] The mixture was kept at 10° C. The size of the amplified PCR product was 948 bp as determined by 1% agarose gel electrophoresis.

[0052] Example 2. Construction of plant recombinant expression vector pCAMBIA1305-OsKTR2 and acquisition of rice transgenic materials

[0053] 1. Construction of OsKTR2 gene expression vector: pCAMBIA1305 plasmid vector was double-digested with SacI and PstI, and the cut linearized expression vector was recovered by gel, and then homologous recombination reaction was carried out with PCR linear product containing OsKTR2. The homologous recombination product was transformed into E. coli DH5α competent cells, and after antibiotic screening, positive single clones were picked and saved. The E. coli plasmid was extracted and sent to the company for sequencing. After correct sequencing, the pCAMBIA1305-OsKTR2 intermediate vector containing Ubiquitin strong promoter was obtained. On this basis, we used EcoRI (GAATTC) and SacI (GAGCTC) to double-digest this vector to obtain a pCAMBIA1305 intermediate vector carrying OsKTR2 gene but removing Ubiquitin promoter, and the enzyme digestion product was recovered by gel. According to the promoter sequence of OsKTR2 gene, a 1964bp sequence was intercepted, and a pair of homologous recombination primers (F2 and R2) for amplifying the promoter of OsKTR2 gene were designed using Primer 5.0 software. EcoRI (GAATTC) and SacI (GAGCTC) restriction site sequences were added to the 5' and 3' ends of the primers, and a sequence on the pCAMBIA1305 plasmid vector was added. Using wild-type Nipponbare rice gDNA as a template, the linear fragment of the target promoter was obtained by PCR technology. After verification, it was homologously recombined with the pCAMBIA1305-OsKTR2 linear vector with the Ubiquitin promoter removed. The homologous recombination product was transformed into Escherichia coli DH5α competent cells. After antibiotic screening, the positive single clones were picked and preserved. The Escherichia coli plasmid was extracted and sent to the company for sequencing. After correct sequencing, the recombinant expression vector pCAMBIA1305-OsKTR2 was obtained, which was transferred into EHA105 Agrobacterium competent cells by electroporation. After antibiotic screening, the positive clones were picked and the plasmid was extracted and transformed back into Escherichia coli. After the extracted plasmid was verified by sequencing, an equal volume of 30% glycerol was added to the Agrobacterium liquid and stored in a -70°C refrigerator for subsequent experiments.

[0054] Upstream primer F2:

[0055] 5'-TATGACCATGATTAC GAATTC TTGTGGGATAGTAAACGAGA-3' (SEQ ID NO.4);

[0056] Downstream primer R1:

[0057] 5'-GGATCCCCGGGTACC GAGCTC GTGTTTGTGTGTGAGCTTGT-3' (SEQ ID NO. 5).

[0058] 2. Obtaining transgenic rice: The Agrobacterium obtained above with the self-promoter overexpression vector of pCAMBIA1305-OsKTR2 was used to infect rice callus tissue, and after co-cultivation (dark culture) for 2.5 days, the bacteria were washed, and the dried callus was transferred to the selection medium containing 500 mg / L carbenicillin and 50 mg / L hygromycin for the first round of selection culture, and cultured at 28°C for 2 weeks. The resistant callus was transferred to the selection medium containing 500 mg / L carbenicillin and 80 mg / L hygromycin for the second round of selection culture, and cultured at 28°C for light until granular resistant callus tissue grew. Pick the bright yellow resistant callus from the same callus and transfer it to a plastic wide-mouth bottle filled with differentiation medium for differentiation culture, wait for differentiation into seedlings (25-30 days), and when the seedlings grow to about 2-3 cm, put them into the rooting medium for seedling strengthening. Pick out the differentiated seedlings from the rooting tube, add an appropriate amount of sterile water, and harden the seedlings for one week. The root agar medium is washed off, and the plants are transplanted into rice nutrient solution for growth and positive seedling identification is performed, and the positive seedlings are transferred to the field for harvesting to obtain T1 generation transgenic seeds. The culture media used are all prior art.

[0059] 3. Identification of overexpression effect of OsKTR2 transgenic rice materials and plant type screening: The T1 generation transgenic seeds of the OsKTR2 overexpression strains obtained above and the wild-type rice seeds of Nipponbare were germinated by water. Select healthy seeds, soak them in 30% sodium hypochlorite solution for 30 minutes, then rinse them with clean water 5 times, place them in a paper cup covered with a 20-mesh nylon mesh, submerge half of the seeds with clean water, and then put them in a 37°C oven for germination for 24-36 hours. Ten days after emergence, all overexpression strains were identified by GUS staining, and positive seedlings were selected and moved to turnover boxes for hydroponics. After culturing with full IRRI nutrient solution for 2 weeks, a part of the seedlings was selected, and the total RNA in the tissues of the OsKTR2 gene overexpression strains and the wild-type Nipponbare wild-type rice was extracted using Trizol reagent, and cDNA was obtained by reverse transcription. Using this as a template, RT-qPCR was used to identify the expression level of the OsKTR2 gene at the transcriptional level. The internal reference and OsKTR2 gene primers used for RT-qPCR are shown in Table 1. Another part of the seedlings were used to screen plant types, and the biomass of all plants was counted ( Figure 1 Among the 9 OsKTR2 overexpressing seedlings tested ( Figure 1 A), the expression of OsKTR2 was upregulated to varying degrees ( Figure 1 B), In addition, the root dry weight of these overexpressing seedlings ( Figure 1 C) and aboveground dry weight ( Figure 1 D) No significant difference with the wild type. Combining the expression level of OsKTR2 and the dry weight of seedlings, we selected three representative strains from these nine overexpressing seedlings and named them Pro1, Pro2 and Pro3 for subsequent physiological experiments.

[0060] Table 1 Primer sequences of OsActin and OsKTR2 used for RT-qPCR

[0061]

[0062] Example 3. Identification of the spatiotemporal expression pattern of rice OsKTR2 and its response to external environmental stress using RT-qPCR technology

[0063] In order to explore the spatiotemporal expression pattern of the OsKTR2 gene, we used the internal reference and OsKTR2 quantitative primers in Example 2, and took samples from different growth stages and different tissues of Nipponbare wild-type rice for RT-qPCR analysis. The specific steps of RT-qPCR are: according to the instructions of the TaKaRa kit, use a 10μL reaction system: SYBR PremixExTaq 5μL, 0.2μL of upstream and downstream primers, 0.2μL of 50×ROX Reference Dye, 2.4μL of ddH2O, and 2μL of diluted cDNA template. The reaction conditions are: 95℃ 5min, 95℃ 5s, 60℃ 30s, 40cycles; 95℃ 15s, 60℃ 1min, 95℃15s. The quantitative results are based on the C T (Threshold Cycle) value is 2 -ΔCt Method calculation analysis. The results are as follows Figure 2 A shows that OsKTR2 is expressed in different tissues of rice throughout the growth period, indicating that OsKTR2 may be widely involved in regulating the expression of downstream genes.

[0064] At the same time, in order to explore the response of OsKTR2 gene to environmental stress, we subjected 10-day-old Nipponbare wild-type rice to different environmental stresses, and took samples from the roots and shoots for RT-qPCR analysis of OsKTR2 expression patterns, and used samples without any treatment as the control group. The results showed that the transcription level of OsKTR2 was affected by low potassium (0.05 mM K + ) Stress-induced upregulation is about 4-10 times ( Figure 3 AB), which was induced by H2O2 (10 mM) to increase by about 3 times ( Figure 3 CD), which was induced by ACC (0.1 mM) to increase by about 2 times ( Figure 3 EF).

[0065] Example 4. Analysis of OsKTR2 subcellular localization

[0066] According to the CDS sequence of OsKTR2, a double restriction site containing BamHI and XbaI and a single restriction site containing BglII were designed, and homologous recombination primers containing the eGFP empty sequence were also designed. Then, the total cDNA sequence of rice was used as a template, and PCR amplification reactions were performed with two pairs of specific homologous recombination of OsKTR2-C and OsKTR2-N (Table 2) to amplify the open reading frame ORF of the OsKTR2 gene. Then, the PCR amplification product was subjected to homologous recombination reaction with the linearized vector pCAMBIA1305.1-GFP (Figure 9) after double restriction digestion and single restriction digestion to obtain the homologous recombination product. After sequencing was correct, the positive strain was stored in a -70°C refrigerator.

[0067] In order to verify whether OsKTR2 has the nuclear localization characteristics of transcription factors, we constructed 2x 35S::OsKTR2::GFP fusion protein and 2x 35S::GFP::OsKTR2 fusion protein recombinant plasmids with 2x 35S cauliflower mosaic virus (CaMV) as promoter. At the same time, we used 2x 35S::GFP fusion protein with 2x 35S cauliflower mosaic virus (CaMV) as promoter as positive control. Then, these three groups of recombinant plasmids were transiently expressed in rice protoplasm, and the subcellular localization of the target protein in rice cells was observed under a laser scanning confocal fluorescence microscope. The results showed that the green fluorescence emitted by both 2x35S::OsKTR2::GFP and 2x 35S::GFP::OsKTR2 fusion proteins can completely overlap with the blue light emitted by the nuclear dye DAPI ( Figure 2 B), while the green fluorescence emitted by the positive control 2x 35S::GFP fusion protein fills the entire cell and cannot completely overlap with the blue light emitted by the nuclear dye DAPI. This indicates that OsKTR2 is located in the cell nucleus and has the characteristics of nuclear localization of transcription factors.

[0068] Table 3 Primer sequences for subcellular localization of OsKTR2

[0069]

[0070] Example 5. Exploring the effect of OsKTR2 expression on rice potassium absorption and accumulation

[0071] In order to explore whether OsKTR2 affects rice's potassium absorption, we cultured 10-day-old Nipponbare wild-type and OsKTR2 overexpressing rice seedlings in low-potassium (0.05mM K) and normal-potassium (1mM K) IRRI nutrient solutions for 4 weeks. During this period, the pH of the nutrient solution was adjusted to 5.1-5.8, and 6 replicates were set for each material. After the treatment, plant samples were collected from the root and aboveground parts, weighed, and the potassium content was determined. The results showed that there was no significant difference in the biomass of the OsKTR2 self-promoter overexpressing material compared with the wild type under low-potassium or normal-potassium conditions ( Figure 4 AD), while the potassium content and total potassium content were significantly higher than those of the wild type ( Figure 4 EH). Under low potassium conditions, the potassium content in the roots and aboveground parts of the OsKTR2 promoter overexpressing materials was higher than that of wild-type rice, about 23%-33% ( Figure 4 E); the total potassium content is about 14%-18% higher than that of the wild type ( Figure 4 G). Under normal potassium conditions, the potassium content in the roots of the OsKTR2 promoter overexpressing material was about 18% higher than that of the wild-type rice, and there was no significant difference in the aboveground part ( Figure 4 F); the total potassium content is about 23%-30% higher than that of the wild type ( Figure 4 H). The above results show that OsKTR2 expression is induced and enhanced by external low potassium stress, thereby promoting the absorption and accumulation of potassium in rice.

[0072] Example 6. Statistics of barrel culture and field phenotypes and main agronomic traits of wild-type and transgenic rice

[0073] In order to study the effects of OsKTR2 on the field phenotype and yield of rice during maturity, we conducted field trials on wild-type and transgenic rice materials. The field trials were conducted at the Pailou Experimental Base of Nanjing Agricultural University. The soil in the experimental base is acidic yellow-brown soil with a pH of about 5.20. The available potassium concentration in the soil was determined to be 120 mg / kg by ammonium acetate extraction (normal K content, and the rest of the nutrients were normal). The field trial was planted in small plots, with each line being 7 x 7 and a total of 49 replicates. During the period, watering, fertilization, and spraying were carried out regularly until the rice was fully mature. The differences in agronomic traits such as plant height, panicle length, total tillering, effective tillering, number of grains per panicle, fruit setting rate, thousand-grain weight, and single plant yield were statistically analyzed ( Figure 5and Tables 3-4), we found that whether under normal potassium supply or low potassium conditions, OsKTR2 self-promoter (Pro) overexpression lines increased rice yield (21-50%); the total tiller number and effective tiller number increased significantly, with an average of 3-6 effective tillers per plant; and the fruit setting rate increased by about 25%. In addition, under normal potassium supply conditions, the number of grains per panicle of OsKTR2 self-promoter (Pro) overexpression lines increased by 11-17% compared with the wild type, while there was no significant difference under low potassium supply conditions. There was no significant difference in plant height, main panicle length and 1000-grain weight compared with the wild type, and the comprehensive single plant yield was significantly increased compared with the wild type. Therefore, we found that whether under normal potassium supply or low potassium conditions, the tillering ability (total tillering and effective tillering) of OsKTR2 self-promoter (Pro) overexpression materials was stronger than that of the wild type, thereby achieving the effect of increasing rice yield.

[0074] Table 3 Statistics of agronomic traits of wild-type and transgenic rice field trials (normally)

[0075] Genotype WT(NB) KTR2PRO1 KTR2PRO2 KTR2PRO3 Plant Height(cm) 90.0±2.07a 90.0±3.92a 85.7±5.05a 88.2±3.43a Total tiller number per plant 16.8±2.12b 21.2±3.77a 20.8±0.45a 20.2±1.94a Effective tiller number per plant 12.3±1.39b 14.6±1.95a 15.7±2.50a 14.3±1.03a Pani cle Length(cm) / Main spike length 21.4±1.23a 22.5±1.49a 20.1±0.91a 21.9±1.27a Grain number per panicle / number of grains per panicle 101.5±5.90b 118.8±8.14a 117.8±5.40a 112.5±7.97b 1000-grain weight(g) 23.4±0.50a 23.0±1.26a 23.0±1.99a 24.0±2.31a Seed setting rate(%) / fruiting rate 73.2±6.87b 82.8±4.00a 82.5±4.36a 83.4±5.26a Yield (g / plant) / yield per plant 14.7±3.24b 21.4±2.81a 19.1±1.20a 21.4±2.91a

[0076] Table 4 Statistics of agronomic traits of wild-type and transgenic rice field trials (low potassium land)

[0077] Genotype WT(NB) KTR2PRO1 KTR2PRO2 KTR2PRO3 Plant Height(cm) 80.1±3.40a 78.0±3.39a 78.3±3.20a 80.5±3.70a Total tiller number per plant 12.9±2.59b 17.8±3.70a 17.7±2.94a 16.8±2.05a Effective tiller number per plant 9.0±1.60b 14.1±1.14a 12.9±3.23a 13.0±1.0a Panicle Length(cm) / Main panicle length 20.9±0.92a 21.0±1.38a 20.2±0.80a 20.4±0.88a Grain number per panicle / number of grains per panicle 90.9±5.96a 90.2±9.01a 88.33±9.58a 90.2±9.65a 1000-grain weight(g) 22.92±1.10a 23.75±1.56a 22.40±1.61a 23.0±2.47a Seed setting rate(%) / fruiting rate 66.9±5.24b 73.6±3.53a 78.6±6.32a 73.7±3.41a Yield (g / plant) 10.7±1.84b 15.8±3.62a 12.9±1.22a 15.3±2.29a

[0078] Although embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that various changes, modifications, substitutions and variations may be made to the embodiments without departing from the principles and spirit of the present invention, and that the scope of the present invention is defined by the appended claims and their equivalents.

Claims

1. Rice ethylene-responsive transcription factor OsKTR2, characterized by The cDNA sequence is shown as SEQ ID NO.

1.

2. A recombinant expression vector, characterized in that Contains the rice ethylene-responsive transcription factor OsKTR2 according to claim 1.

3. The recombinant expression vector according to claim 2, characterized in that The starting vector is pCAMBIA1305 vector.

4. The recombinant expression vector according to claim 3, characterized in that The recombinant expression vector replaces the Ubiquitin promoter of the pCAMBIA1305 vector with the promoter of the rice ethylene response transcription factor OsKTR2.

5. The recombinant expression vector according to claim 3, characterized in that The recombinant expression vector is obtained by inserting the rice ethylene response transcription factor OsKTR2 described in claim 1 into the SacI and PstI restriction sites of the starting vector pCAMBIA1305, then removing the Ubiquitin promoter with EcoRI and SacI enzymes and inserting the OsKTR2 gene's own promoter.

6. The use of the rice ethylene response transcription factor OsKTR2 according to claim 1, characterized in that: Select one or more of the following: (1) Increase the total number of tillers, effective tillers, and fruit setting rate of crops; (2) Increase rice yield; (3) Promote rice's absorption of potassium.

7. Use of the rice ethylene-responsive transcription factor OsKTR2 according to claim 1 in constructing transgenic rice, characterized in that: The transgenic rice has any one or more of the following traits: (1) Increase the total number of tillers, effective tillers, and fruit setting rate of crops; (2) Increase rice yield; (3) Promote rice's absorption of potassium.

8. Use of the recombinant expression vector according to any one of claims 2 to 5, characterized in that: Select one or more of the following: (1) Increase the total number of tillers, effective tillers, and fruit setting rate of crops; (2) Increase rice yield; (3) Promote rice's absorption of potassium.

9. Use of the recombinant expression vector according to any one of claims 2 to 5 in constructing transgenic rice, characterized in that: The transgenic rice has any one or more of the following traits: (1) Increase the total number of tillers, effective tillers, and fruit setting rate of crops; (2) Increase rice yield; (3) Promote rice's absorption of potassium.

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