KR101819448B1 - Gene implicated in drought stress tolerance and drought stress resistant plants using the same - Google Patents

Abstract

The present invention relates to a dry stress resistance related gene and a dry stress resistant plant produced using the same. Since the dry stress resistance gene of the present invention participates in the resistance to dry stress of the plant by participating in the pore opening and closing process, a plant having increased resistance to dry stress can be obtained by preparing an overexpressed transgenic plant using the plant.

Description

[0001] The present invention relates to a dry stress tolerance-related gene and a dry stress-tolerant plant using the same,

The present invention relates to a dry stress resistance related gene and a dry stress resistant plant produced using the same.

Because higher plants are inaccessible during their lifetimes, they face environmental stresses such as drought, high salt, heavy metals, cold weather, thermal shock and ozone. These abiotic stresses are limiting factors for the growth and development of crops, and it has been confirmed that over half of the possible production is lost due to abiotic stress due to the loss of crop production due to abiotic environmental stress. In addition, recent rapid changes in the environment have been recorded in local areas.

Of these abiotic stresses, water deficit is the most serious environmental factor considered to be a major cause of crop production decline. Worldwide, water consumption continues to increase, and the availability of clean water can be an important issue not only for humans but also for higher plants. Plants operate a variety of defense strategies, such as regulating signaling pathways or inducing stress-responsive genes to increase resistance to short- or long-term water shortages. The mechanism of cellular or genetic defense against such water stress is well known (Shinozaki and Yamaguchi-shinozaki, 2007). Yet, knowledge of the biological functions of stress-related genes involved in stress tolerance and sensitivity in higher plants is still lacking.

On the other hand, GMO (Genetic Modified Organism) is a genetically modified organism that is produced by introducing DNA isolated from a living organism. It is not only resistant to various diseases and environmental stress according to the development of biotechnology, Fragrance, nutritional value, pigment, etc. have been applied to the development of increased crops. In addition, industrial and pharmacological substances that can be used for the prevention and treatment of diseases of the human body can be used as a means for mass production with ease. Therefore, a functional study of stress response genes is important to increase crop productivity. Since 1996, the organization has been annually providing a database of GMO crop cultivation areas around the world as official data for the International Service for the Acquisition of Agri-Biotech Applications (ISAAA / www.isaaa.org). According to this, since 1996, the cultivation of GMO crops has been steadily increasing worldwide and commercialization has been actively carried out.

Therefore, it is necessary to develop stress-related genes that are involved in tolerance and sensitivity to dry stress, and to produce GMOs using the stress-related genes.

Accordingly, the inventors of the present invention have conducted studies to discover genes capable of enhancing tolerance to drying stress of a plant. As a result, the present inventors have completed the present invention by confirming that the Arabidopsis thaliana AFBA1 gene is involved in the dry stress resistance of plants.

It is an object of the present invention to provide a dry stress resistance gene comprising the nucleotide sequence of SEQ ID NO: 1.

It is still another object of the present invention to provide a vector containing the gene, and a plant transformed cell transformed with the vector.

It is still another object of the present invention to provide a method for producing a dry stress-resistant plant including a step of preparing a vector containing the gene and introducing the vector into the plant, and a method for increasing the dry stress resistance of the plant.

It is still another object of the present invention to provide a composition for increasing the dry stress resistance of a plant comprising the vector.

In order to achieve the above object, the present invention relates to a dry stress-resistance gene comprising the nucleotide sequence of SEQ ID NO: 1AFBA1of to provide.

In addition, the present invention provides a protein consisting of the amino acid sequence of SEQ ID NO: 2.

The present invention also provides a vector comprising the gene.

In addition, the present invention provides a dry stress-tolerant plant transformed cell which is transformed with said vector to increase the expression of AFBA1.

The present invention also provides a method for producing a vector comprising the steps of: 1) preparing a vector comprising a gene consisting of the nucleotide sequence of SEQ ID NO: 1; 2) preparing a plant transformed cell transformed with the vector prepared in the step 1); And 3) preparing a plant with the plant transformed cell prepared in the step 2).

The present invention also provides a method for producing a vector comprising the steps of: 1) preparing a vector comprising a gene consisting of the nucleotide sequence of SEQ ID NO: 1; And 2) introducing the vector prepared in the step 1) into a plant to increase the expression of AFBA1, thereby providing a method for increasing the dry stress resistance of the plant.

The present invention also provides a composition for increasing dry stress resistance of a plant comprising the vector.

In addition, the present invention provides a composition for identification of a dry stress-resistant plant comprising an agent for measuring the expression of the AFBA1 gene.

In addition, the present invention provides a kit for identification of a dry stress-resistant plant comprising an agent for measuring the expression of the AFBA1 gene.

Since the dry stress resistance gene of the present invention participates in resistance to drying stress by participating in the pore opening and closing process, a plant having increased resistance to dry stress can be obtained by using the transgenic plant to produce an overexpressed transgenic plant.

Figure 1 shows the effect of AFBA1 on ABB1 (ABA hormone, salt, dry stress) And the expression pattern of the gene was confirmed by real-time PCR.
Figure 2 shows AFBA1 This is a schematic diagram for producing an overexpressed gene.
Fig. 3 is a diagram showing the base sequence of AFBA1 (At1g12490) gene derived from Arabidopsis thaliana.
4 shows AFBA1 In the overexpressed gene, AFBA1 And the amount of expression of the gene was confirmed by RT-PCR.
Figure 5 shows AFBA1 This is a schematic diagram for producing a gene expression inhibiting mutant.
FIG. 6 shows AFBA1 In the gene expression inhibition mutant, AFBA1 And the amount of expression of the gene was confirmed by RT-PCR.
Figure 7 shows AFBA1 < RTI ID = 0.0 > Overexpressing genes and AFBA1 The phenotype of the gene expression inhibition mutant was confirmed.
Figure 8 shows AFBA1 Overexpressing genes and AFBA1 FIG. 5 is a graph showing the results of measurement of water loss in the leaves of the gene expression-inhibiting mutants over time.
Figure 9 shows that when treated with the ABA hormone or untreated, AFBA1 Fig. 5 shows the result of microscopic analysis of pore opening and closing of mutant of AFBA1 gene expression inhibiting mutant.
Figure 10 shows that AFBA1 Overexpressing genes and AFBA1 FIG. 5 is a graph showing the results of analyzing pore sizes of gene expression inhibiting mutants. FIG.

Hereinafter, the present invention will be described in detail.

The present invention provides a dry stress resistance gene AFBA1 consisting of the nucleotide sequence of SEQ ID NO: 1.

The AFBA1 of the present invention Gene (ABA insensitive FBA motif-containing protein 1) is the At1g12490 gene of Arabidopsis-derived, DNA sequence of the gene is AFBA1 915bp. Preferably, the gene sequence is composed of the nucleotide sequence of SEQ ID NO: 1.

Variants of the above base sequences are also included within the scope of the present invention. Specifically, the gene includes a nucleotide sequence having at least 70% homology, more preferably at least 80% homology, even more preferably at least 90% homology, and most preferably at least 95% homology with the nucleotide sequence of SEQ ID NO: 1 can do. "% Of sequence homology to polynucleotides" is ascertained by comparing the comparison region with two optimally aligned sequences, and a portion of the polynucleotide sequence in the comparison region is the reference sequence for the optimal alignment of the two sequences (I. E., A gap) relative to the < / RTI >

The gene is expressed in plants, preferably Arabidopsis thaliana .

The expression level of the AFBA1 gene is increased under abiotic stresses such as ABA hormone, salt, and dry stress.

The term "dry stress" in the present invention means a condition in which water use of the plant is restricted.

The term "dry stress resistance" in the present invention means a phenotype that resumes normal growth under a condition that water loss from leaves is low in dry stress condition and water is supplied after drying stress.

In addition, the present invention provides a protein consisting of the amino acid sequence of SEQ ID NO: 2. The protein can be coded by the dry stress resistance gene AFBA1 . The present invention includes a protein consisting of the amino acid sequence of SEQ ID NO: 2 and functional equivalents of said protein. Is at least 70%, preferably at least 80%, more preferably at least 90%, even more preferably at least 70%, more preferably at least 90%, more preferably at least 90% Refers to a protein having substantially the same physiological activity as a protein having an amino acid sequence represented by SEQ ID NO: 2, having a sequence homology of 95% or more. In addition, the protein of the present invention includes not only a protein having a native amino acid sequence thereof but also mutants thereof, within the scope of the present invention. By "variant" is meant a protein having a sequence that differs by deletion, insertion, non-conservative or conservative substitution, or a combination thereof, with the amino acid sequence of AFBA1 and one or more amino acid residues.

The present invention also provides a vector comprising the dry stress resistance gene AFBA1 . The term "vector" in the present invention is used to refer to a DNA fragment (s), a nucleic acid molecule, which is transferred into a cell. The vector replicates the DNA and can be independently regenerated in the host cell. Vector refers to a recombinant DNA molecule comprising a desired coding sequence and a suitable nucleic acid sequence essential for expressing a coding sequence operably linked in a particular host organism and includes a promoter of the invention and a target operably linked to the promoter Viruses or other mediators known in the art to which a gene sequence encoding a protein can be inserted or introduced. In one embodiment of the present invention, pCAMBIA1300, a binary vector for plant transformation, was used.

For purposes of the present invention, "operably linked" means that a suitable molecule is linked in such a way as to enable gene expression when binding to an expression control sequence.

In the present invention, the term "binary vector" refers to a plasmid having a LB (left order) and RB (right border) necessary for movement in a Ti (tumor inducible) plasmid and a plasmid having a gene necessary for transferring a target nucleotide Vector illustration.

The base sequence of the promoter and the gene of the vector may be operatively linked to an expression control sequence and the operably linked gene sequence and expression control sequence may be operably linked to a single marker comprising a selectable marker and a replication origin Expression vector. An " expression control sequence "means a DNA sequence that regulates the expression of a polynucleotide sequence operably linked to a particular host cell. Such regulatory sequences include promoters for conducting transcription, any operator sequences for regulating transcription, sequences encoding suitable mRNA ribosome binding sites, and sequences controlling the termination of transcription and translation.

In a preferred embodiment of the present invention, the promoter may be a promoter for plant expression such as CMV (Cauliflower Mosaic Virus) 35S promoter, CMV 19S promoter, Agrobacterium tumefaciens Ti plasmid, nos synthase promoter, ocs (octopine synthase) promoter and mas (mannopine synthase) promoter, and a known promoter.

The vector may preferably comprise one or more selectable markers. The marker is typically a nucleic acid sequence having a property that can be selected by a chemical method, and includes all genes capable of distinguishing a transformed cell from a non-transformed cell. Examples include herbicide resistance genes such as glyphosate or phosphinotricin (phosphinotricin), kanamycin, G418, Bleomycin, hygromycin, chloramphenicol, , But are not limited thereto.

In addition, the present invention provides a dry stress-tolerant plant transformed cell which is transformed with said vector to increase the expression of AFBA1.

Transformation of a plant means any method of transferring DNA to a plant. Such transformation methods do not necessarily have a regeneration and / or tissue culture period. Transformation of plant species is now common for plant species, including both terminal plants as well as dicotyledonous plants. In principle, any transformation method can be used to introduce the hybrid DNA according to the present invention into suitable progenitor cells. For example, calcium chloride (CaCl ₂ ) and heat shock method, particle gun bombardment, silicon carbide whiskers, sonication, electroporation and PEG (Polyethylenglycol) And a precipitation method based on the method. A preferred method according to the present invention comprises Agrobacterium mediated DNA transfer, and the transforming Agrobacterium of the present invention may be any gene that is suitable for the expression of a nucleotide sequence. In one embodiment of the present invention, Agrobacterium tumefaciens GV3101 was used as Agrobacterium strain for agar.

The "plant cell" used for the transformation of a plant can be any plant cell. Plant cells may be any of the cultured cells, cultured tissue, cultured or whole plant, preferably cultured cells, cultured tissue or culture medium, and more preferably cultured cells.

The term "plant tissue" is intended to encompass a variety of cells used in differentiated or undifferentiated plant tissues such as, but not limited to, roots, stems, leaves, pollen, seeds, Protoplasts, shoots and callus tissue. The plant tissue may be in planta or may be in an organ culture, tissue culture or cell culture.

In the present invention, the term "plant" includes food crops including rice, wheat, barley, corn, soybean, potato, red bean, oats and millet; Vegetable crops including Arabidopsis, cabbage, radish, red pepper, strawberry, tomato, watermelon, cucumber, cabbage, melon, squash, onions, onions and carrots; Special crops including ginseng, tobacco, cotton, sesame seed, sugar cane, beet, perilla, peanut, and rapeseed; Fruit trees including apple trees, pears, jujube trees, peaches, sheep grapes, grapes, citrus fruits, persimmons, plums, apricots, and bananas; Roses, gladiolus, gerberas, carnations, chrysanthemums, lilies, tulips; And feed crops including rice grass, red clover, orchardgrass, alfalfa, tall fescue, perennial rice, and the like. However, the present invention is not limited thereto.

The present invention also provides a method for producing a vector comprising the steps of: 1) preparing a vector comprising a gene consisting of the nucleotide sequence of SEQ ID NO: 1; 2) preparing a plant transformed cell transformed with the vector prepared in the step 1); And 3) preparing a plant with the plant transformed cell prepared in the step 2); And a method for producing a dry stress-tolerant plant.

In the present invention, the vector used for inserting the dry stress-resistance gene AFBA1 consisting of the nucleotide sequence of SEQ ID NO: 1 may be any general expression vector for plant transformation. In one embodiment of the present invention, Gt; pCAMBIA1300 < / RTI > was used to produce a plant overexpressing AFBA1 .

The present invention also provides a method for producing a vector comprising the steps of: 1) preparing a vector comprising a gene consisting of the nucleotide sequence of SEQ ID NO: 1; And 2) introducing the vector prepared in the step 1) into a plant to increase the expression of AFBA1, thereby providing a method for increasing the dry stress resistance of the plant.

(Methods of Enzymology, Vol. 153, (1987)) in order to produce plant transformed cells and transgenic plants of the present invention. Plasmids can be transformed by inserting the exogenous polynucleotide into a carrier such as a plasmid, a virus, etc., and Agrobacterium bacteria can be used as a mediator (Chilton et al. Cell 11: 263: 271 (1977)), Direct exogenous polynucleotides can be introduced into plant cells to transform plants (Lorz et al. MoI Genet. 199: 178-182; (1985)). For example, when a vector not containing a T-DNA region is used, electroporation, microparticle bombardment, and polyethylene glycol-mediated uptake may be used.

Generally, a method widely used for transforming plants is a method of infecting plant cells or seeds with Agrobacterium tumefaciens transformed with an exogenous polynucleotide. One of ordinary skill in the art can cultivate or plant transformed plant cells or seeds under suitable known conditions to develop into plants.

As used herein, the term "plant (sieve) " is understood to mean not only mature plants but also plant cells, plant tissues and plant seeds that develop into mature plants.

Selection of plant transformed cells can be carried out by exposing the transformed cultures to a selection agent (e.g., metabolic inhibitor, antibiotic and herbicide). Plant cells that stably contain a marker gene that is transformed and conferring selectative resistance are grown and divided in the above cultures. Exemplary labels include, but are not limited to, the hygromycin phosphotransferase gene, the glycophosphate tolerance gene, and the neomycin phosphotransferase (nptII) system. Methods for the development or regeneration of plants from plant protoplasts or from various expansions are well known in the art. The development or regeneration of plants containing foreign genes introduced by Agrobacterium can be accomplished according to methods known in the art.

The present invention also provides a method for producing a vector comprising the steps of: 1) preparing a vector comprising a gene consisting of the nucleotide sequence of SEQ ID NO: 1; And 2) introducing the vector prepared in the step 1) into a plant to increase the expression of AFBA1, thereby providing a method for increasing the dry stress resistance of the plant.

According to the method of increasing the dry stress resistance of the plant of the present invention, the sensitivity of the pore opening and closing can be increased by overexpression of the AFBA1 gene, so that it is possible to grow even under the dry stress condition, It is effective.

In addition, the method of increasing the dry stress resistance of the plant of the present invention has an advantage that the dry stress resistance can be increased without affecting other functions of the plant.

The present invention also provides a composition for increasing dry stress resistance of a plant comprising a vector comprising a dry stress-resistance gene AFBA1 .

The vector contained in the composition for increasing the dry stress resistance of the plant is as described above.

In addition, the present invention provides a composition for identification of a dry stress-resistant plant comprising an agent for measuring the expression of the AFBA1 gene.

In the present invention, the term "agent for measuring the expression of AFBA1 gene" means a preparation used for confirming the expression of AFBA1 contained in a sample, preferably RT-PCR, competitive RT- (PCR), real-time RT-PCR, Western blotting and the like, the probe or primer capable of specifically binding to the target gene can be used. But is not limited to.

The term "primer" refers to a nucleic acid sequence having a short free 3 'hydroxyl group, capable of forming base pairs with a complementary template and functioning as a starting point for template strand copying. ≪ / RTI > The primers can initiate DNA synthesis in the presence of reagents and four different nucleoside triphosphates for polymerization reactions (i. E., DNA polymerase or reverse transcriptase) at appropriate buffer solutions and temperatures.

The term "probe" refers to a nucleic acid fragment such as RNA or DNA corresponding to a short period of a few nucleotides or a few hundreds of nucleotides capable of specifically binding with a gene or an mRNA. The oligonucleotide probe, short-chain DNA single stranded DNA probes, double stranded DNA probes, RNA probes, and the like, and can be labeled for easier detection.

Was AFBA1 an increased expression by a dry stress in the specific embodiment of the present invention, since confirmed that the resistance to drought stress increase in the over-expression element which overexpressing AFBA1, when using the agent measuring the AFBA1 level of the present invention dry It can be seen that stress tolerant plants can be identified effectively.

In addition, the present invention provides a kit for identification of a dry stress-resistant plant comprising an agent for measuring the expression of the AFBA1 gene.

The kit may further comprise one or more other components, solutions or devices suitable for analyzing the amount of gene expression, gene expression level or expression pattern, or the method for analyzing the amount or presence pattern of the protein in addition to nucleic acid or antibody . For example, when the kit is a kit for detecting the expression level or expression pattern of a gene, it may be a kit containing essential components necessary for performing RT-PCR. The RT-PCR kit may include a biomarker gene (DNTPs), enzymes such as Taq-polymerase and reverse transcriptase, DNase, RNase inhibitor (RNase inhibitor), and the like, depending on the specific embodiment, DEPC water, sterilized water, primer pairs specific for genes used as a quantitative control, and the like. On the other hand, if the kit is a kit for detecting the amount or presence pattern of a protein, the kit may be, for example, a kit containing essential components necessary for performing an ELISA, (E.g., an antibody conjugated with an antibody) and its substrate, and an antibody specific for a quantitative control protein, and the like, for example, . According to a specific embodiment, the kit may comprise a DNA microarray or a protein microarray.

Terms not otherwise defined herein have meanings as commonly used in the art to which the present invention belongs.

Hereinafter, the present invention will be described in detail with reference to examples. However, the following examples are illustrative of the present invention and are not intended to limit the scope of the present invention.

Example  One: Abiotic  Stress Treatment AFBA1 Identification of gene expression pattern

For a variety of abiotic stress treatments of ABA hormone, salt, and dry environment, two weeks old Arabidopsis thaliana was grown in MS medium (Mursahige and Shoog medium). For the ABA stress treatment, plants grown as above were treated with 100 [mu] M ABA hormone, treated with 300 mM of salt by spraying method on the above-grown plants for salt stress treatment, Plants grown in the medium were picked up and placed on a filter paper for drying and securing the plants in time zones. RNA was extracted from the obtained plants to obtain a sample, and AFBA1 The expression pattern of the gene was confirmed by real-time PCR and the results are shown in FIG.

As shown in FIG. 1, in the stress-treated group, AFBA1 The expression of the gene was increased up to 3 hours and gradually decreased. In the case of dry stress, it was confirmed that the dry stress was strongly increased until 3 hours.

Therefore, it was confirmed that the expression of AFBA1 gene was induced by various abiotic stresses. Especially, in case of dry stress, the expression of AFBA1 gene was increased in proportion to the time of stress application.

Example  2: AFBA1 Gene separation and Cloning

Arabidopsis a primer set comprising a BamH sequence at the terminal 'Xba sequence and 3 at the terminal, 5 to the mold (template) for a total of gDNA (genomic DNA) of thaliana ecotype Columbia) (forward primer:. TGAGGATCCGATTAAACTCTGGAAATAACAAAGA; reverse primer: CATTCTAGAATGGTCTCCAGAAGTCGTGAGGAA SEQ ID NO: 3 , 4) amplified the At1g12490 ( AFBA1 ) gene by PCR to obtain an amplified product of 915 bp.

The PCR product was digested with Xba and BamH restriction enzymes, and a binary vector (CaMV) 35S promoter was inserted between a left boarder and a right boarder. ) PCAMBIA1300 vector, and a schematic diagram of such cloning was shown in Fig.

As a result of sequencing the prepared clone, it was confirmed that the correct base sequence was cloned. AFBA1 The sequence of the gene is shown in Fig.

Example  3: AFBA1  gene Overexpression  And expression inhibition mutants

The vector prepared in Example 2 was introduced into Agrobacterium GV3101 by a freeze-thaw method (An, G. 1987, Methods in Enzymology). Arabidopsis thaliana was transfected with transformed Agrobacterium in flower dipping method (Clough and Bent, 1998, The Plant Journal). The transformed Arabidopsis thaliana was screened through three generations to secure the T3 line.

RT-PCR was performed using AFBA1 primer (forward primer: ATCTGATTATTFCCAATGTATTATGGTGTAAT; reverse primer: GCCTTGGTAAAAAAAGACTCTGGATATATCATAAT, SEQ ID NOS: 5 and 6) in order to confirm the expression level of AFBA1 in the selected AFBA1 overexpressing transformants. Respectively. ELF4a was used as an internal control group.

As shown in FIG. 4, AFBA1 overexpression ( AFBA1- Ox line) was found to increase the expression of AFBA1 as compared with that of wild type (WT).

On the other hand, the AFBA1 expression-suppressing mutant (afba1) was obtained from the SIGNAL Salk Institute Genomuc Analysis Laboratory (http://signal.salk.edu/) by using a mutant Arabidopsis seed (seed number: Salk_133974C) ≪ / RTI > Fig. 5 shows the production scheme of the AFBA1 expression-inhibiting mutant.

RT-PCR was performed using AFBA1 primer (the same as SEQ ID NO: 3) to confirm that the obtained AFBA1 expression-suppressing mutant actually inhibited the expression of AFBA1 , and the expression was compared. Respectively.

As shown in Fig. 6, it was confirmed that the expression was inhibited in the AFBA1 expression inhibiting mutant.

Example  4: AFBA1  gene Overexpression  And Sensitivity to Dry Stress of Expression Inhibiting Mutants

In order to measure the sensitivity of the AFBA1 gene overexpression construct and the inhibitory mutation construct constructed in Example 3 to the dry stress, the seeds of wild type, AFBA1 expression inhibiting mutant, and AFBA1 overexpressing strain were grown for 2 weeks in soil , And the degree of tolerance to the dry stress after water was not given for 10 days and then water was again confirmed by phenotype observation. The results are shown in Fig.

As shown in FIG. 7, AFBA1 expression-suppressing mutants showed almost phenotypic dying after drying stress (Drought) compared to wild type, whereas AFBA1 overexpressed bodies were healthy and green compared with wild type, and leaf tautness was maintained .

In the re-watering condition in which the water was rehydrated , AFBA1 expression-suppressing mutants showed almost no survival phenotype, whereas AFBA1 overexpressing agents were continuously growing.

Therefore, AFBA1 The gene is resistant to dry stress, and it can be seen that the dry stress is constantly growing after the drought condition and drought.

To assess the response to further drying stress, the water loss percentage in the leaves of the plants was measured. Leaves with similar size, growth period, and position in each wild-type, AFBA1 expression inhibiting mutant, and AFBA1 overexpression were isolated and the leaf weight reduction was measured according to the known method (Sang et al., 2001). The results are shown in Fig.

As shown in Fig. 8, under AFBA1 overexpression, the weight loss of AFBA1 overexpressing mutants was slower than that of wild type leaves, and the weight loss of AFBA1 inhibitory mutant leaves was slightly faster than that of wild type leaves Respectively.

Thus, the weight reduction effect decreases appear in AFBA1 overexpressed body is because reducing the water loss of AFBA1 overexpressed body leaves AFBA1 The genes show dry stress resistance by reducing water loss in plant leaves.

Example  5: Opening and closing the pore stomatal  aperture measurement

AFBA1 To confirm the effect of the gene on the pore opening, wild type grown in soil for 4 weeks, AFBA1 expression inhibiting mutant, AFBA1 After removing the leaves from the overexpressed cells, they were immersed in a pore-opening solution (5 mM MES-KOH, 20 mM KCl, pH 6.15) for 2 hours and then transferred to a solution containing ABA hormone . After that, only the epithelial layer of each plant leaf was peeled off, and the degree of opening and closing of the pores was observed through a microscope, and the result was shown in a graph, which is shown in FIG.

As shown in Fig. 9, before treatment with ABA hormone, wild type, AFBA1 expression inhibiting mutant, AFBA1 The pores of overexpressed bodies were well open. On the other hand, compared with wild type after ABA hormone treatment AFBA1 The pore of the expression-inhibiting mutant was still open and AFBA1 The pores of the overexpressed leaves were closed as in the wild type.

This phenomenon is quantified and the results are shown in Fig.

As shown in FIG. 10, the size of the AFBA1- inhibiting mutant pore size was about 22% larger than that of the wild type, and the AFBA1 overexpression pore size was about 5% smaller than that of the wild type .

Therefore, AFBA1 expression-inhibiting mutants decreased the susceptibility to pore opening and closing, resulting in plants dying quickly when subjected to dry stress, while AFBA1 Overexpressed cells are very sensitive to the pore opening and closing activity of the guard cells, so that the pores of the guard cells close quickly even in the absence of water, so that the plants show a phenotype in which growth is maintained. The data show that the AFBA1 gene regulates the water content of the plant by regulating the pore opening and closing of the guard cells in the dry stress condition, which leads to resistance to the drying stress.

Although the present invention has been described in terms of the preferred embodiments mentioned above, it is possible to make various modifications and variations without departing from the spirit and scope of the invention. It is also to be understood that the appended claims are intended to cover such modifications and changes as fall within the scope of the invention.

<110> Korea University Research and Business Foundation <120> Gene implicated in drought stress tolerance and drought stress          resistant plants using the same <130> 1_58P <160> 6 <170> Kopatentin 2.0 <210> 1 <211> 915 <212> DNA <213> arabidopsis thaliana <400> 1 atggtctcca gaagtcgtga ggattatttt aacccggatt taaaacatct aactacactg 60 gtgttgggtt catcatcatc ggtcaccatc cctacacctt gggagaagga caaggagaag 120 gagaaggaga aggagaagga ggacgaagaa ttcttcttag tttcttttga tagctgtgac 180 ggtctcgttt gtctctacaa atactggaaa tctggttatg tggtcaatcc caccactaga 240 tggtatcgcc ctcttcctct ctctcaatta caacaactcc ttatcagctt aggtcgcagt 300 gtcttcgagc ttggatacac agtctgtgat attggattcg gtaaagacaa aatcacgggc 360 acatacaaac ccgtttggtt atacaattct ttagaaattg gccttgaaaa cgctactaca 420 tgcgaggttt tcgactttaa caccaacgct tggagatatg tttctccgac cgctccctat 480 cgggaggaaa ccaaaattct atctttcgat cttcacactg aaacctttcg agtggtctct 540 aaagctcctt ttaccaatgt aaaagctttt gatatcgtca tgtgcaatct cggcaaccgc 600 ttgtgcgttt ccgagaagaa ctggcccaac caagtgatat ggtcattcaa ttcaggcaac 660 aagacatggc acaaaatgtt ttccattaat cttgatgtaa cttctcattg gtttggtaat 720 cacatagccg cggtcatgcc actagcactt ttttatgaga agaagaagaa gaagaagttg 780 ttgttttatt gtcgtgtgcg aagtcgaaca ttgatggtat atgatcccga aaccgaatca 840 tatgatgttg ctttcaatga ttactccatc ggttatcctc tttgttattt ccagagttta 900 atctctattt cataa 915 <210> 2 <211> 304 <212> PRT <213> arabidopsis thaliana <400> 2 Met Val Ser Ser Arg Glu Asp Tyr Phe Asn Pro Asp Leu Lys His   1 5 10 15 Leu Thr Thr Leu Val Leu Gly Ser Ser Ser Ser Val Thr Ile Pro Thr              20 25 30 Pro Trp Glu Lys Asp Lys Glu Lys Glu Lys Glu Lys Glu Lys Glu Asp          35 40 45 Glu Glu Phe Leu Val Ser Phe Asp Ser Cys Asp Gly Leu Val Cys      50 55 60 Leu Tyr Lys Tyr Trp Lys Ser Gly Tyr Val Val Asn Pro Thr Thr Arg  65 70 75 80 Trp Tyr Arg Pro Leu Pro Leu Ser Gln Leu Gln Gln Leu Leu Ile Ser                  85 90 95 Leu Gly Arg Ser Val Phe Glu Leu Gly Tyr Thr Val Cys Asp Ile Gly             100 105 110 Phe Gly Lys Asp Lys Ile Thr Gly Thr Tyr Lys Pro Val Trp Leu Tyr         115 120 125 Asn Ser Leu Glu Ile Gly Leu Glu Asn Ala Thr Thr Cys Glu Val Phe     130 135 140 Asp Phe Asn Thr Asn Ala Trp Arg Tyr Val Ser Pro Thr Ala Pro Tyr 145 150 155 160 Arg Glu Glu Thr Lys Ile Leu Ser Phe Asp Leu His Thr Glu Thr Phe                 165 170 175 Arg Val Val Ser Lys Ala Pro Phe Thr Asn Val Lys Ala Phe Asp Ile             180 185 190 Val Met Cys Asn Leu Gly Asn Arg Leu Cys Val Ser Glu Lys Asn Trp         195 200 205 Pro Asn Gln Val Ile Trp Ser Phe Asn Ser Gly Asn Lys Thr Trp His     210 215 220 Lys Met Phe Ser Ile Asn Leu Asp Val Thr Ser His Trp Phe Gly Asn 225 230 235 240 His Ile Ala Ala Val Met Pro Leu Ala Leu Phe Tyr Glu Lys Lys Lys                 245 250 255 Lys Lys Lys Leu Leu Phe Tyr Cys Arg Val Val Arg Ser Thr Leu Met             260 265 270 Val Tyr Asp Pro Glu Thr Glu Ser Tyr Asp Val Ala Phe Asn Asp Tyr         275 280 285 Ser Ile Gly Tyr Pro Leu Cys Tyr Phe Gln Ser Leu Ile Ser Ile Ser     290 295 300 <210> 3 <211> 34 <212> DNA <213> Artificial Sequence <220> <223> AFBA1 PCR forward primer <400> 3 tgaggatccg attaaactct ggaaataaca aaga 34 <210> 4 <211> 33 <212> DNA <213> Artificial Sequence <220> <223> AFBA1 PCR reverse primer <400> 4 cattctagaa tggtctccag aagtcgtgag gaa 33 <210> 5 <211> 32 <212> DNA <213> Artificial Sequence <220> <223> AFBA1 RT PCR forward primer <400> 5 atctgattat tgccaatgta ttatggtgta at 32 <210> 6 <211> 35 <212> DNA <213> Artificial Sequence <220> <223> AFBA1 RT PCR reverse primer <400> 6 gccttggtaa aaaaagactc tggatatatc assign 35

Claims (11)

A composition for inducing dry stress resistance of a plant comprising a dry stress-resistance gene AFBA1 consisting of the nucleotide sequence of SEQ ID NO: 1.
A composition for inducing dry stress resistance of a plant comprising a protein consisting of the amino acid sequence of SEQ ID NO: 2.
delete A dry stress-tolerant plant transformation cell, which is transformed with a vector comprising the dry stress-resistance gene AFBA1 consisting of the nucleotide sequence of SEQ ID NO: 1, whereby the expression of AFBA1 is increased.
5. The method according to claim 4, wherein the plant is selected from the group consisting of food crops including rice, wheat, barley, corn, soybeans, potatoes, red beans, oats and millet; Vegetable crops including Arabidopsis, cabbage, radish, red pepper, strawberry, tomato, watermelon, cucumber, cabbage, melon, squash, onions, onions and carrots; Special crops including ginseng, tobacco, cotton, sesame seed, sugar cane, beet, perilla, peanut, and rapeseed; Fruit trees including apple trees, pears, jujube trees, peaches, sheep grapes, grapes, citrus fruits, persimmons, plums, apricots, and bananas; Roses, gladiolus, gerberas, carnations, chrysanthemums, lilies, tulips; And feed crops including rice grapes, red clover, orchardgrass, alfalfa, tall fescue, and penny al raglas.
1) preparing a vector comprising a gene consisting of the nucleotide sequence of SEQ ID NO: 1;
2) preparing a plant transformed cell transformed with the vector prepared in the step 1); And
3) preparing a plant with the plant transformed cell prepared in the step 2); &Lt; / RTI &gt;
1) preparing a vector comprising a gene consisting of the nucleotide sequence of SEQ ID NO: 1; And
2) increasing the expression of AFBA1 by introducing the vector prepared in the step 1) into the plant, thereby increasing the dry stress resistance of the plant.
8. The method of claim 7, wherein said method is capable of increasing dry stress resistance without affecting other functions of the plant.
delete 1. A composition for identification of a dry stress-tolerant plant comprising an agent for measuring the expression of an AFBA1 gene comprising the nucleotide sequence of SEQ ID NO: 1.
1. A kit for identification of a dry stress-tolerant plant comprising an agent for measuring the expression of an AFBA1 gene comprising the nucleotide sequence of SEQ ID NO: 1.

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