Coding gene for enhancing iron deficiency stress tolerance of plants and application

Abstract

The invention is applicable to the technical field of bioengineering, and provides a coding gene for enhancing iron deficiency stress tolerance of plants and application. According to the soybean GmMYB14 transcription factor disclosed by the invention, the problem that the growth of soybeans is limited in an iron-deficient environment, particularly calcareous soil and soda saline alkali soil, is solved by excavating and applying the encoding gene (the nucleotide sequence is shown as SEQ ID No.1) of the soybean GmMYB14 transcription factor. Experiments prove that after the gene is over-expressed in arabidopsis thaliana and soybean, the tolerance of plants to iron deficiency stress can be remarkably enhanced. The invention provides a key gene resource for molecular breeding: the GmMYB14 effectively promotes the absorption and utilization of plants on iron by regulating and controlling the metabolic pathway of phenylpropane, not only can improve the planting adaptability of soybeans in low-iron soil, but also can improve the yield and quality of the soybeans, provides powerful technical support for enlarging the planting area of the soybeans in iron-deficient soil regions, and has a wide application prospect. The method is of great significance in relieving insufficient soybean productivity in China and improving the self-sufficiency rate.

Description

Technical Field

[0001] The present invention belongs to the field of bioengineering technology, and in particular relates to a coding gene for enhancing plant tolerance to iron deficiency stress and its application. Background Art

[0002] Iron (Fe), the most required trace element for soybean growth and development, is not only involved in multiple biological processes such as respiration and photosynthesis, but is also a key component of nitrogenase and leghemoglobin in soybean root nodules, making it a crucial factor influencing nodule nitrogen-fixing activity. Despite the high total iron content in the Earth's crust, iron bioavailability is generally low in well-aerated calcareous or soda-alkali soils, leading to frequent iron deficiency in plants. Iron deficiency can cause yellowing of new leaves and decreased photosynthetic rates in crops from the seedling stage. It can also reduce the formation and activity of underground root nodules, and in severe cases, can be lethal. This significantly impacts soybean yield and quality, resulting in significant economic losses for agricultural production. Therefore, iron deficiency is a key factor limiting crop yield in agricultural production. It is reported that over one-third of arable soils worldwide are at risk of iron deficiency. The traditional approach to addressing soybean iron deficiency is to apply iron fertilizers. While this approach can temporarily alleviate iron deficiency symptoms, it has drawbacks such as low utilization efficiency and high environmental risks. With the development of molecular biology, molecular breeding technology has provided a revolutionary approach to overcoming the bottleneck of iron deficiency stress. This technology fundamentally improves plants' adaptability to iron-deficient environments by precisely targeting and regulating their own iron absorption and utilization mechanisms.

[0003] As an important dual-purpose crop for both food, oil, and feed, soybeans are an indispensable strategic resource in the global agricultural economy. In recent years, with the improvement of people's living standards, the demand for soybeans has continued to increase. However, my country's soybean production capacity is insufficient and mainly relies on imports. Therefore, it is urgent to expand the soybean planting area and improve its yield and quality. With the deepening of genomic research in crops such as soybean, transcription factor engineering has become a core strategy for improving crop stress tolerance. Among them, MYB transcription factors have been shown to synergistically regulate multiple abiotic stress response pathways and have become an important resource for molecular breeding. The application of molecular breeding can not only significantly reduce the dependence of agricultural production on chemical fertilizers, but also quickly and targetedly improve soybean traits. Therefore, molecular design breeding targeting the MYB network will become the only way to solve the problems of cultivated land alkalinization and iron nutritional disorders.

[0004] MYB transcription factors (especially the R2R3-MYB subfamily) are the core regulators of phenylpropanoid metabolism. They precisely control the spatiotemporal expression of branch pathways such as lignin and flavonoids by binding to cis-elements in the promoters of target genes. In soybean, at least several abiotic stress-induced R2R3-MYB transcription factor genes have been identified (such as GmMYB111, GmMYB76, and GmMYB117). These genes show specific expression patterns under adversities such as salt stress and cold tolerance. As the core hub of plant secondary metabolism, the phenylpropanoid metabolic pathway and its products play multiple physiological roles in the response to iron deficiency stress. On the one hand, this pathway can promote the root system's absorption of Fe by synthesizing coumarins as iron-reducing chelators. 3+ On the other hand, it can affect root architecture by regulating lignin deposition, altering nutrient acquisition capacity. Studies have shown that under iron deficiency, the phenylpropanoid metabolic pathway in soybean roots is rapidly activated, and differential expression of genes for key phenylpropanoid biosynthesis enzymes (such as PAL and 4CL) can be detected even after short periods of iron deficiency. At the metabolite level, the phenylpropanoid metabolic pathway in iron-efficient soybeans is continuously upregulated, and its products chelate iron ions in the soil and maintain their mobility within the body.

[0005] In view of the current problems of low soybean production capacity, low self-sufficiency rate, and high dependence on foreign countries in my country, issues such as increasing soybean yield and quality and expanding planting area have attracted much attention. Traditional iron fertilizer application methods have problems such as low utilization rate and high environmental risks. Although they can temporarily alleviate iron deficiency symptoms, they are not a long-term solution. While molecular breeding technology provides a direction for solving problems such as iron deficiency stress, it is still insufficient in discovering genes that can simultaneously improve soybean alkali tolerance, iron deficiency tolerance, and soybean iron content. To this end, the present invention proposes a coding gene and application for enhancing plant tolerance to iron deficiency stress. Summary of the Invention

[0006] The purpose of the present invention is to provide a coding gene for enhancing plant tolerance to iron deficiency stress and its application, aiming to solve the problems raised in the above background technology.

[0007] The purpose of the present invention is achieved through the following technical solutions: A gene encoding a soybean GmMYB14 transcription factor GmMYB14 Application of the invention in enhancing plant tolerance to iron deficiency stress, the encoding gene GmMYB14 The nucleotide sequence is shown in SEQ ID No.1.

[0008] Furthermore, the plant is soybean or Arabidopsis thaliana.

[0009] Furthermore, the application is to enhance the planting ability of soybeans in low-iron soil, and the low-iron soil includes calcareous soil and soda saline-alkali soil.

[0010] Furthermore, the coding gene GmMYB14 The DNA sequence includes a DNA sequence that has more than 90% homology with the sequence shown in SEQ ID NO.1 and encodes a protein with the same function.

[0011] A sort of pro35S::GmMYB14 Overexpression vector, comprising a coding gene with a nucleotide sequence as shown in SEQ ID NO.1 GmMYB14 , and the pro35S::GmMYB14 The overexpression vector was selected from pCAMBIA3301-35S-GmMYB14 or PTF101-35S-GmMYB14.

[0012] A method for enhancing plant tolerance to iron deficiency stress by using a recombinant bacterium comprising the pro35S::GmMYB14 The overexpression vector is selected from Escherichia coli DH5α or Agrobacterium tumefaciens GV3101; wherein, Escherichia coli DH5α is used for cloning and preservation of the overexpression vector, and Agrobacterium tumefaciens GV3101 is used for mediating the transformation of the overexpression vector into plants.

[0013] To construct the above pro35S::GmMYB14 Primer pairs for overexpression vectors, including: 3301-MYB14-F and 3301-MYB14-R, as shown in SEQ ID No. 3 and SEQ ID No. 4, respectively; or 101-MYB14-F and 101-MYB14-R, as shown in SEQ ID No. 5 and SEQ ID No. 6, respectively.

[0014] A method for enhancing plant tolerance to iron deficiency stress, comprising: The nucleotide sequence of the coding gene is shown in SEQ ID NO. GmMYB14 Through the above pro35S::GmMYB14 The overexpression vector is transformed into plant cells to make the coding gene GmMYB14 Overexpression in plants; The transformed cells are cultivated to obtain complete plants, and single-copy homozygous plants are selected according to Mendel's laws of inheritance.

[0015] Furthermore, the plant is soybean or Arabidopsis thaliana; Arabidopsis thaliana is transformed by floral dip method; soybean is transformed by cotyledonary node transformation method mediated by Agrobacterium GV3101.

[0016] Compared with the prior art, the present invention has the following beneficial effects: The present invention mines and applies the coding gene of soybean GmMYB14 transcription factor GmMYB14(systematically named Glyma.15g259400), specifically addresses the core problem of soybean growth restriction in iron-deficient environments, especially calcareous soils and soda-salt-alkali soils. Experiments have confirmed that overexpression of this gene in Arabidopsis and soybean can significantly enhance the plant's tolerance to iron deficiency stress. This invention overcomes the drawbacks of traditional iron fertilizer application, such as low utilization rate and high environmental risks, and provides a key gene resource for molecular breeding: GmMYB14 effectively promotes the absorption and utilization of iron by plants by regulating the phenylpropanoid metabolic pathway. It can not only improve the planting adaptability of soybeans in low-iron soils, but also improve their yield and quality, providing strong technical support for expanding the planting area of ​​soybeans in iron-deficient soil areas. This achievement has laid the foundation for cultivating iron-deficiency-tolerant crops, reducing agriculture's dependence on chemical fertilizers, and promoting the development of precision breeding. It is of great significance to alleviating my country's insufficient soybean production capacity and increasing self-sufficiency. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] Figure 1 for GmMYB14 Agarose gel electrophoresis diagram of gene PCR amplification and gene structure diagram; A is GmMYB14 Agarose gel electrophoresis of gene PCR amplification; B is GmMYB14 Diagram of the structure of a gene.

[0018] Figure 2 for GmMYB14 Schematic diagram of the construction of gene overexpression vectors; A is a schematic diagram of the construction of the Arabidopsis overexpression vector pCAMBIA3301-35S-GmMYB14; B is a schematic diagram of the construction of the soybean overexpression vector PTF101-35S-GmMYB14.

[0019] Figure 3 Arabidopsis wild type Col-0 and pro35s::GmMYB14 (OE-GmMYB14) The growth of each overexpression line under different iron treatment conditions for 7 days; A is the growth phenotype; B is the Arabidopsis heterologous overexpression line GmMYB14 expression level; C is root length; D is total chlorophyll content; E is relative fluorescence value of root exudates; F is aboveground biomass; G is root biomass; H is root iron content; I is aboveground iron content.

[0020] Figure 4 The soybean wild type Williams 82 and pro35s::GmMYB14 (OE-GmMYB14) Two overexpression lines treated under iron deficiency conditions for 7 days; A is the phenotype; B is the soybean overexpression line GmMYB14 expression level; C is SPAD value; D is total chlorophyll content; E is relative fluorescence value of root exudates; F is aboveground biomass; G is root biomass; H is aboveground iron content. DETAILED DESCRIPTION

[0021] In order to have a clearer understanding of the technical features, objectives and beneficial effects of the present invention, the technical solution of the present invention is now described in detail below, but it should not be understood as limiting the scope of implementation of the present invention.

[0022] The following is a detailed description of the specific implementation of the present invention in conjunction with specific examples. Unless otherwise specified, the experimental methods in the examples are all conventional methods.

[0023] Example 1: (1) By analyzing the transcriptome data of soybeans treated with iron deficiency stress, we discovered a key transcription factor GmMYB14 (whose encoding gene GmMYB14 The system is named Glyma.15g259400 ) and obtained relevant sequence information from the National Center for Biotechnology Information (https: / / www.ncbi.nlm.nih.gov). Among them, the amino acid sequence of the transcription factor GmMYB14 is shown in SEQ ID No. 2, and its encoding gene GmMYB14 The nucleotide sequence is shown in SEQ ID No. 1, and the structure diagram of the gene is drawn at the same time, as shown in Figure 1 As shown in B. In the experiment, total RNA was extracted from the roots of soybean wild type Williams 82 after iron deficiency treatment, and the gene GmMYB14 cDNA. PCR reaction system refers to Table 1, reaction procedure refers to Table 2, and primers used refer to Table 3. PCR amplification products were identified by agarose gel electrophoresis. Figure 1 As shown in A, a clear band can be seen at approximately 1146 bp, and the sequencing results of the amplified product are consistent with SEQ ID No. 1.

[0024] SEQ ID No. 1:

[0025] SEQ ID No. 2: MGKGRAPCCDKTQVKRGPWSPAEDLKLIAFIQKYGHENWRALPKQAGLLRCGKSCRLRWINYLRPDVKRGNFTPEEEETIIRLHKALGNKWSKIASGLPGRTDNEIKNVWNTHLKRLAPKKVSESSADESKPESSITSSSSSSSESFFSNERPNSPKTTTPSNNEFNDQAEKLSVEDKMDKDSEKQLSNE LVNITEDPKESSVSLSSVESNIVNTSQIVEHKPEEEQLASPLPYLGHYDVGSILEEVDKPNHLIEIPWEPDYDFWKLLDDDNSLGSFQSNEVQLGEFPAIQNIILGEEGVHDAEAKKWSHDFENEFGVVGGIKESSKDHFLPKNYAVEPEMDHTQAFDFDDIDMARPESELDFGYIQLWPSCPQNNNTSL*.

[0026] Table 1 PCR reaction system for vector construction

[0027] Table 2 PCR reaction program for target fragment amplification

[0028] Table 3 Primers for expression vector construction

[0029] (2) Construction of Arabidopsis GmMYB14 genetic pro35S::GmMYB14 The overexpression vector pCAMBIA3301-35S-GmMYB14, the vector construction diagram is shown in Figure 2 Middle A; Building soybean GmMYB14 genetic pro35S::GmMYB14 The overexpression vector PTF101-35S-GmMYB14, the vector construction diagram is shown in Figure 2 B. The specific method is as follows: GmMYB14 When the gene is constructed into a plant expression vector, a 35S enhancer promoter (used in this example) or other enhancer promoters or inducible promoters can be added before the transcription start nucleotide. Using the cDNA from the roots of soybean wild-type Williams 82 treated with iron deficiency as a template, the high-fidelity enzyme PrimeSTAR HS from Takara was used to amplify the gene. GmMYB14The amplified product was identified by agarose gel electrophoresis (see Figure 1 A), the gel was cut and recovered according to the instructions of the TIANGEN general gel recovery kit. The recovered cDNA fragments were constructed into the 35S promoter downstream of the linearized pCAMBIA3301-35S-NOS vector treated with SpeI restriction endonuclease single enzyme digestion and the linearized PTF101-35S-NOS vector treated with XbaI and SacI restriction endonucleases. The connection process was carried out according to the instructions of 2×Seamless Cloning Mix. The ligation product was transformed into Escherichia coli DH5α competent cells and incubated upside down at 37°C for 16h; single clones were selected and cultured at 37°C and 200rpm for 12h, and sequenced and compared after identification as positive clones by bacterial solution PCR. An equal volume of 50% glycerol was added to the bacterial solution that was successfully sequenced and stored at -80°C for a long time. The corresponding pro35S::GmMYB14 The overexpression vectors were named pCAMBIA3301-35S-GmMYB14 and PTF101-35S-GmMYB14, respectively.

[0030] Plasmid acquisition: Take 100µL of the above-preserved bacterial solution and add it to 100ml LB liquid medium containing the corresponding antibiotic. The corresponding antibiotic for pCAMBIA3301-35S-GmMYB14 is kanamycin (Kana), and the corresponding antibiotic for PTF101-35S-GmMYB14 is spectinomycin (Spec); culture at 37℃, 200rpm for 12h, and extract the plasmid according to the instructions of the TIANGEN plasmid extraction kit to obtain GmMYB14 genetic pro35S::GmMYB14 Overexpression vector.

[0031] (3) The above-mentioned pro35S::GmMYB14 The overexpression vector was transformed into Agrobacterium tumefaciens GV3101 (Agrobacterium-mediated transformation was used in this example; transformation can also be accomplished using conventional biological methods such as Ti plasmids, plant virus vectors, direct DNA transformation, microinjection, and electroporation). The specific method is as follows: 40 µL of competent Agrobacterium cells (aseptically) were slowly thawed on ice, followed by the addition of 2 µL of the extracted plasmid (approximately 200 ng). The cells were then placed on ice for 5 minutes, frozen in liquid nitrogen for 5 minutes, and heat-shocked at 37°C for 5 minutes. The cells were then added with 1 mL of LB liquid medium and incubated at 200 rpm and 28°C for 2 hours. The incubated culture was then spread onto LB plates containing the corresponding antibiotics (rifampicin and kanamycin) and incubated inverted at 28°C for 2 days. Single colonies were selected for colony PCR identification. If the PCR-positive culture was identified, an equal volume of 50% glycerol was added and the culture was stored at -80°C for long-term storage.

[0032] (4) Arabidopsis thaliana and soybean were transformed using the pCAMBIA3301-35S-GmMYB14 and PTF101-35S-GmMYB14 expression vectors, respectively.

[0033] For Arabidopsis transformation, the floral dip method was used. The detailed method was as follows: Agrobacterium containing the pCAMBIA3301-35S-GmMYB14 expression vector was first inoculated into 5 mL LB (kanamycin Kana and rifampicin Rif) liquid medium at a ratio of 1:10, and cultured overnight at 28°C and 200 rpm (12-16 h); then 3 mL of the Agrobacterium culture solution was transferred into 300 mL LB (containing the corresponding antibiotics and Rif for the plasmid) liquid medium at a ratio of 1:10, and cultured overnight at 28°C and 200 rpm (12-16 h) to achieve an OD of 0. 600 The bacterial suspension was then poured into a 400mL centrifuge bottle and centrifuged at 4000 rpm for 15 minutes at room temperature (Beckman Avanti JXN-26 high-speed centrifuge). The supernatant was removed and the cells were harvested. The suspension was resuspended in 250mL of infiltration solution and the Arabidopsis thaliana plants to be transformed were placed in this solution for 1 minute. The soaked plants were placed in a transfer box (or tray) in the dark for 24 hours before removal and continued cultivation in the greenhouse. The first-generation seeds were screened for resistance on ½MS solid medium containing glufosinate-ammonium resistance (to facilitate identification and screening of transgenic plants). Positive plants were identified and harvested. Positive plants that met Mendel's law of segregation at a ratio of 3:1 were further propagated and individually harvested for homozygous identification. Gene expression levels were used to confirm the presence of overexpressing plants.

[0034] For soybean transformation, the PTF101-35S-GmMYB14 expression vector was transformed into soybeans using an Agrobacterium-mediated genetic transformation system at the cotyledonary node. Positive plants were screened and propagated in a laboratory greenhouse. The following year, the plants were propagated in a transgenic seedling facility. Positive plants were identified by foliar spraying with 25 μg / mL of glufosinate. Before conducting the iron deficiency experiment, each seedling was sprayed with glufosinate and expression levels were verified. Positive plants were then confirmed for subsequent experiments.

[0035] (5) Conduct molecular testing on transgenic lines to verify GmMYB14 The expression of genes in each strain was detected by qRT-PCR for Arabidopsis pCAMBIA3301-35S-GmMYB14 transgenic lines. GmMYB14 The relative expression of genes, as shown in Figure 3 As shown in B, wild-type Arabidopsis (Col-0) GmMYB14 The gene expression level was extremely low, while in the homozygous overexpression Arabidopsis lines #10, #13, #14 and #19, GmMYB14The gene expression level was high, and the expression level of strain #10 was the highest. GmMYB14 The relative expression of genes, as shown in Figure 4 As shown in B, wild-type soybean (WT) GmMYB14 The gene expression level was extremely low, while in the homozygous soybean lines #13 and #17, GmMYB14 The gene expression level was high, and the expression level of strain #13 was the highest. GmMYB14 The gene can be successfully and efficiently transcribed and expressed in Arabidopsis overexpression lines and soybean overexpression lines.

[0036] (6) Phenotypic observation, physiological indicators and iron content detection of transgenic lines.

[0037] Overexpression GmMYB14 The Arabidopsis lines showed stronger tolerance to iron deficiency and iron utilization than Col-0 under iron deficiency treatment (½MS, 0 μM Fe) (see Figure 3 ), compared with Col-0, overexpressing Arabidopsis has more developed root system, milder iron deficiency chlorosis symptoms, less inhibition of aboveground growth, longer root length, higher total chlorophyll content, more fluorescent phenolic root secretions, higher aboveground biomass, higher root biomass, higher root iron content, higher aboveground iron content and other phenotypes. Specifically, overexpression GmMYB14 Alleviates the aboveground chlorosis symptoms and root growth inhibition caused by iron deficiency in Arabidopsis (see Figure 3 Middle A), average root length of wild-type Arabidopsis after iron deficiency treatment (see Figure 3 The root length of the overexpressing Arabidopsis homozygous lines #10, #13, #14, and #19 were 4.03 cm, 3.69 cm, 3.55 cm, and 3.65 cm, respectively. The root length of the transgenic plants treated with iron deficiency increased by 7.1%-21.6% compared with the wild-type plants. Figure 3 The total chlorophyll content of the transgenic plants treated with iron deficiency increased by 12.6%-57.5% compared to that of the wild-type plants. Figure 3The fluorescence value of the root exudates of the transgenic plants treated with iron deficiency increased by 35.9%-121.7% compared with the wild-type plants. Figure 3 The iron content of the transgenic plants was 1.03 mg / plant, while that of the overexpressing Arabidopsis homozygous lines #10, #13, #14, and #19 were 1.36 mg / plant, 1.21 mg / plant, 1.33 mg / plant, and 1.32 mg / plant, respectively. The aboveground biomass (fresh weight) of the transgenic plants treated with iron deficiency increased by 18.2%-32.2% compared with that of the wild-type plants. The average root biomass of the wild-type Arabidopsis after iron deficiency treatment (see Figure 3 The iron content of the transgenic plants treated with iron deficiency was 7.2%-50.0% higher than that of the wild-type plants. Figure 3 The iron content of the transgenic plants in the wild-type was 9.0%-194.6% higher than that of the wild-type plants. Figure 3 The iron content in the aboveground parts of the transgenic plants treated with iron deficiency was 43.5%-101.1% higher than that of the wild-type plants.

[0038] Overexpression GmMYB14 Soybean lines grown in low-iron hydroponic conditions (modified ¼ Hoagland's hydroponic nutrient solution containing 1 μM Fe-EDTA) showed greater tolerance to iron deficiency than wild-type (WT) soybeans (see Figure 4 ), which is reflected in the phenotypes of milder top leaf chlorosis, higher SPAD value, higher total chlorophyll content, more fluorescent phenolic root secretions, and higher aboveground iron content. Specifically, under iron deficiency conditions, the aboveground growth of the overexpressing soybean lines is less inhibited than that of the wild type, and the yellowing of the top leaves of the wild type soybean is more obvious (see Figure 4A), SPAD values ​​of wild-type soybean after iron deficiency treatment (see Figure 4 The SPAD value of the transgenic plants treated with iron deficiency was 45.0%-120.0% higher than that of wild-type soybean. Figure 4 The total chlorophyll content of the transgenic plants treated with iron deficiency was 170.8%-332.6% higher than that of the wild-type soybean. Figure 4 The fluorescence value of the root secretions of the transgenic soybean plants treated with iron deficiency increased by 830%-3800% compared with that of the wild-type soybean. Figure 4 The aboveground iron content of transgenic soybean plants treated with iron deficiency was 30.9%-44.5% higher than that of wild-type soybean. Under normal growth conditions, the aboveground biomass of wild-type soybean (see Figure 4 F) was 12.60g, while the overexpression homozygous lines #13 and #17 were 4.21g and 7.63g respectively; after iron deficiency treatment, the aboveground biomass of wild-type soybean was 6.83g, while the overexpression homozygous lines #13 and #17 were 3.67g and 5.33g respectively. Under normal growth conditions, the root biomass of wild-type soybean (see Figure 4 The aboveground biomass of the wild-type soybean was 2.11 g, while that of the overexpressing homozygous lines #13 and #17 was 0.72 g and 1.82 g, respectively. These data indicate that the transgenic soybean plants had less biomass than the wild-type soybean. Under normal growth conditions, the aboveground biomass of the transgenic plants was 33.4%-60.6% of that of the wild-type soybean, and the root biomass was 32.7%-62.6%. However, after iron deficiency treatment, the aboveground biomass of the transgenic plants was 53.7%-78.1% of that of the wild-type soybean, and the root biomass was 34.3%-86.1%. This indicates that iron deficiency stress has a lesser effect on biomass changes in the transgenic plants than in the wild-type soybean.

[0039] The present invention is constructed by genetic engineering technology pro35S::GmMYB14The overexpression vectors (pCAMBIA3301-35S-GmMYB14 and PTF101-35S-GmMYB14) were respectively transformed into Arabidopsis wild-type Col-0 plants by floral dipping method and into soybean wild-type Williams 82 plants by Agrobacterium-mediated cotyledonary node transformation method. GmMYB14 The gene is overexpressed in wild-type plants, making the plants tolerant to iron deficiency. GmMYB14 The transgenic strains of the plant showed significantly enhanced tolerance to iron deficiency. Compared with the wild type, overexpression of this gene not only increased the biomass, root length and leaf chlorophyll content of the plant under iron deficiency conditions, but also significantly increased the iron content of the plant. Based on the functional verification of this gene in Arabidopsis and soybean, it is speculated that it may have similar effects of enhancing iron deficiency tolerance in other monocotyledons (such as rice, wheat, corn, etc.) and dicotyledons (such as rapeseed, tobacco, cucumber, tomato, poplar, lawn grass, alfalfa, etc.). The present invention is of great significance for the analysis of the mechanism of soybean tolerance to iron deficiency stress, and provides genetic resources and technical support for the expansion of soybean cultivation in low-iron soils (especially calcareous soils and soda saline-alkali soils).

[0040] The above are only preferred embodiments of the present invention. It should be pointed out that for those skilled in the art, several variations and improvements can be made without departing from the concept of the present invention. These should also be regarded as the scope of protection of the present invention. These will not affect the effect of the implementation of the present invention and the practicality of the patent.

Claims

1. A gene encoding a soybean GmMYB14 transcription factor GmMYB14 The application of the invention in enhancing the tolerance of plants to iron deficiency stress is characterized in that: The coding gene GmMYB14 The nucleotide sequence is shown in SEQ ID No.

1.

2. The use according to claim 1, characterized in that The plant is soybean or Arabidopsis thaliana.

3. The use according to claim 1, characterized in that The application is to enhance the planting ability of soybean in low-iron soil, and the low-iron soil includes calcareous soil and soda saline-alkali soil.

4. The use according to claim 1, characterized in that The coding gene GmMYB14 The DNA sequence includes a DNA sequence that has more than 90% homology with the sequence shown in SEQ ID NO.1 and encodes a protein with the same function.

5. A pro35S::GmMYB14 The overexpression vector is characterized in that Contains the coding gene shown in SEQ ID NO.1 GmMYB14 , and the pro35S::GmMYB14 The overexpression vector was selected from pCAMBIA3301-35S-GmMYB14 or PTF101-35S-GmMYB14.

6. Use of a recombinant bacterium in enhancing plant tolerance to iron deficiency stress, characterized in that: The recombinant bacteria comprises the pro35S::GmMYB14 The overexpression vector is selected from Escherichia coli DH5α or Agrobacterium tumefaciens GV3101; wherein, Escherichia coli DH5α is used for cloning and preservation of the overexpression vector, and Agrobacterium tumefaciens GV3101 is used for mediating the transformation of the overexpression vector into plants.

7. For constructing the method according to claim 5 pro35S::GmMYB14 The primer pair of the overexpression vector is characterized in that: include: 3301-MYB14-F and 3301-MYB14-R, as shown in SEQ ID No. 3 and SEQ ID No. 4, respectively; or 101-MYB14-F and 101-MYB14-R, as shown in SEQ ID No. 5 and SEQ ID No. 6, respectively.

8. A method for enhancing plant tolerance to iron deficiency stress, characterized in that: include: The nucleotide sequence of the coding gene is shown in SEQ ID NO. GmMYB14 According to claim 5 pro35S:: GmMYB14 The overexpression vector is transformed into plant cells to make the coding gene GmMYB14 Overexpression in plants; The transformed cells are cultivated to obtain complete plants, and single-copy homozygous plants are selected according to Mendel's laws of inheritance.

9. The method according to claim 8, characterized in that The plant is soybean or Arabidopsis thaliana; the floral dip method is used to transform Arabidopsis thaliana; and the cotyledon node transformation method mediated by Agrobacterium GV3101 is used to transform soybean.

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