KR100930593B1 - Sweet Potato Root Derived Protein and Its Gene Encoding It - Google Patents

Abstract

The present invention relates to swDREB1 derived from sweet potato root, and more particularly, to a dry stress-inducible DREB protein derived from dried sweet potato root and a gene encoding the same. Since the swDREB1 gene of the present invention is strongly induced by various stresses such as dryness, low temperature, and salt in the roots of sweet potato plants, it may be usefully used to develop an environmental stress resistant plant suitable for a disadvantageous area.

SwDREB1, Stress, RT-PCR

Description

SwDREB1 protein originated from a root of Ipomoea batatas and gene coding the same}

The present invention is swDREB1 derived from sweet potato root The present invention relates to a protein and a gene encoding the same, and more particularly, to a swDREB1 protein, which is a transcription factor derived from sweet potato root, which is highly expressed by dry, low temperature, and various compounds, and a gene encoding the same.

Sweet potato ( Ipomoea batatas L. Lam) can be grown on relatively poor lands and has a yield of about 22 tons per hectare, which is a representative root crop used for food and livestock feed. In particular, about 70% of the buildings are made of starch, which has been used as a raw material for alcoholic beverages for a long time, and has recently been re-examined as an environmentally friendly alternative energy crop for bioethanol production.

Recently, due to the rapid industrialization and population increase, environmental and food problems on the global scale have been raised, and the development of environmentally resistant crops that grow well even in areas where desertification, high seas and cold regions are disadvantageous has been actively conducted. In particular, in order to develop industrial sweet potatoes suitable for the disadvantaged area, research on environmentally resistant transformed sweet potatoes by molecular breeding has been attempted (Lim et al. Mol Breeding 19, 227-239, 2007).

In order to develop environmentally resistant sweet potatoes to improve the early growth of seedlings in poor soil such as dry and cold, the development of genes that are highly expressed in the sweet potato roots (fibrous root) is urgently needed. In dry stress conditions due to lack of moisture, the development of thread roots is thought to be important for tuber growth. In addition, since the roots develop and grow into tubers, it is thought that the development of genes that are highly expressed in the roots is important when producing useful materials from the tubers. However, no genes have been reported that specifically express the sweet potato roots under various stress conditions including drying. However, genes encoding the sporamin protein, which is highly expressed in tubers of sweet potatoes, have long been isolated and studied (Hattori et al. Plant Mol Biol 14, 595-604, 1990).

In this regard, the isolation of genes that are strongly expressed in sweet potato roots under stress conditions may be useful for developing transgenic sweet potatoes producing environmentally resistant plants and various useful materials. In other words, metabolic engineering using advanced biotechnologies, especially when growing sweet potato varieties such as dry land, reclaimed land, and high sea areas, producing industrially useful materials such as starch, functional protein, and so on. It is expected to be able to produce eco-friendly bioenergy without affecting.

DREB (DRE-binding protein) transcription factor of the present invention is a DRE (Dehydration responsive element: TACCGACAT of a promoter of the rd29A , rd15 / cor47 , kin1 , cor6 .6 / kin2 gene, which is a dry, low-temperature, salt stress inducer in Arabidopsis; ) Is a transcription factor that binds to cis activity regulators (Wang et al. Plant Mol Biol 28, 605-617, 1995; Iwasaki et al. Plant Physiol 115, 1287-1294, 1997). Recently, DRE cis activity regulators were identified in promoters of 16 dry-induced genes in Arabidopsis, and these genes were found to bind DREB1 or DREB2 family transcription factors (Seki et al. Plant J 31, 279-292, 2002). In the Arabidopsis, where genomic information was fully identified, 55 DREB transcription factors were identified, and their expression was regulated by signaling mechanisms dependent or independent of the plant hormone ABA under stress conditions. However, no DREB gene has been reported in sweet potatoes (Stockinger et al. Proc Natl Acad Sci USA 94, 1035-1040, 1997; Liu et al. Plant Cell 10, 1391-1406, 1998; Sakuma et al. Biochem Biophys Res Commun 290, 998-1009, 2002).

Thus, the present inventors have completed the present invention by confirming that the swDREB1 gene isolated from dried sweet potato silt is specifically expressed in silt root under various stress conditions including drying.

An object of the present invention is to provide a swDREB1 protein derived from dried sweet potato root and a gene encoding the same.

In order to achieve the above object, the present invention provides a swDREB1 protein and its amino acid sequence derived from sweet potato root.

The present invention also provides a polynucleotide encoding the swDREB1 protein.

In addition, the present invention provides an expression vector comprising the polynucleotide.

The present invention also provides a transformant transformed with an expression vector comprising the polynucleotide.

In addition, the present invention provides a method for producing a transformed plant having a stress resistance comprising the polynucleotide.

The present invention also provides a method for the high expression of foreign proteins in the roots.

Hereinafter, the present invention will be described in detail.

The present invention provides swDREB1 protein and its amino acid sequence derived from sweet potato root.

The swDREB1 protein of the present invention is characterized by having an amino acid sequence set forth in SEQ ID NO: 2 .

The swDREB1 protein of the present invention is composed of 257 amino acids, and the NLS, AP2 / EREBP site and acidification site of the C terminus of the general DREB protein structure are well conserved in the swDREB1 protein amino acid sequence. In particular, in the swDREB1 gene of the present invention, DSAWRL and LWSF, which are specific sequences of the DREB1 series of the Arabidopsis DREB series, were conserved (see FIG. 1).

In addition, pepper ( Capsicum ) when the coding portion of the swDREB1 of the present invention was irradiated with blast (BlastX) in annuum) CaCBF1B, CaDREBLP1, tomato (Lycopersicon In LeCBF1, AtDREB1D / CBF4 and amino acid levels of Arabidopsis thaliana (Arabidopsis thaliana) of esculentum) showed a homology of 61-67% (see Fig. 2a and 2b).

The present invention also provides a polynucleotide encoding the swDREB1 protein.

The polynucleotide encodes a protein having an amino acid sequence set forth in SEQ ID NO: 2, and preferably has a nucleotide sequence set forth in SEQ ID NO: 1 .

The swDREB1 cDNA gene of the present invention has a total length of 1,206 bp, an untranslated region (UTR) at the 5 'end of 132 bp, an untranslated region at the 3' end of 300 bp and a coding portion of 774 bp (consisting of 257 amino acids). It consists of (refer FIG. 1).

The swDREB1 gene of the present invention was strongly expressed in the tuberous root and stem. It was also expressed in fibrous root but very weakly in leaf and thick pigmented root (see FIG. 3).

In addition, the swDREB1 gene of the present invention showed the strongest expression level at 1 hour after drying treatment in the root root tissue, and maintained the strong expression level until 16 hours (see FIG. 4A). On the other hand, no increase or decrease in the expression level of swDREB1 was observed in the leaf tissues. In addition, swDREB1 was not observed to increase or decrease the expression of the dry stress-related plant hormone Apsis acid (see FIG. 4B). SwDREB1 gene was strongly increased in leaf and root expression by low temperature (4 ℃) of various temperature stress (see Figure 4c). Various compound stresses, such as sodium chloride, methyl viologen, and cadmium treatment, also showed strong expression increases in leaves and roots, respectively (see FIG. 4D).

Therefore swDREB1 has a strong responsiveness to various stresses, including dry stress The gene is a transcription factor that regulates the expression of various stress genes and can be very useful for developing plants that are resistant to various stresses including dry stress.

In addition, the present invention provides an expression vector comprising the polynucleotide.

The gene included in the expression vector of the present invention includes a polynucleotide encoding swDREB1 of the present invention, and preferably includes a polynucleotide having a nucleotide sequence represented by SEQ ID NO: 1 . As the vector used to insert the gene, it is preferable to select and use any one of a pKBS1-1 vector, a pBI101 vector, and a pCAMBIA vector, and any expression vector for general plant transformation may be used.

The present invention also provides a transformed host cell transformed with an expression vector comprising the polynucleotide.

The transformed host cell includes transformed microorganisms, animal cells, plant cells, transformed animals or plants, and cultured cells derived from them.

In addition, the present invention

1) preparing an expression vector comprising a gene construct consisting of a polynucleotide of SEQ ID NO: 1 ;

2) introducing the expression vector into a host plant cell;

3) selecting a transgenic plant cell into which the expression vector is introduced;

4) preparing a callus by inducing germination of the transgenic plant cells; And

5) It provides a method of producing a transgenic plant having a stress resistance comprising the step of inducing differentiation of the callus.

DRE-binding protein (DREB) is a nucleotide sequence set forth in SEQ ID NO: 1 encoding a dry, low temperature and transcription factor that binds to the dehydration responsive element (TACCGACAT) cis activity regulator of the promoter of various compound stress inducing genes. Transgenic plants transformed with an expression vector comprising a polynucleotide having a resistance to the stress.

In addition, the scope of the present invention also includes a plant that is a progeny or clone of the stress-resistant transformed plant and seeds, fruit, ear, tuber, tuber, main, callus or protoplast of the plant.

In addition, the present invention

1) preparing an expression vector comprising a gene construct consisting of a polynucleotide of SEQ ID NO: 1 and a polynucleotide encoding a foreign protein;

2) introducing the expression vector into a host plant cell;

3) selecting a transgenic plant cell into which the expression vector is introduced;

4) preparing a callus by inducing germination of the transgenic plant cells;

5) preparing a transgenic plant by inducing differentiation of the callus; And

6) provides a method for high expression of the foreign protein in the root comprising the step of culturing the transgenic plant.

Preferably, the plant cell is preferably a monocotyledonous plant or a dicotyledonous plant, but is not limited thereto.

The monocotyledonous plants include Alismataceae, Hydrocharitaceae, Juncaginaceae, Schuchzeriaceae, Pomomotontonaceae, Najadaceae, Zosteraceae, Liliaceae, Haemodoraceae, Agavaceae, Amaryllidaceae, Dioscoreaceae, Pontederiaceae, Iridaceae, Burmanniaceae, Juncaceae, Commelinaceae ), Eriocaulaceae, Flower Family (Grapeaceae, Gramineae, Poaceae), Araceae, Lemnaceae, Spaganiaceae, Typhaceae, Cycloaceae, Cyperaceae, Pachoaceae ( Musaceae), Zingiberaceae, Cannaaceae, Orchidaceae (Orchidaceae) is preferably, but not limited to.

The dicotyledonous plants are Asteraceae (Dolaceae, Diapensiaceae), Asteraceae (Clethraceae), Pyrolaceae, Ericaceae, Myrsinaceae, Primaceae (Primulaceae), Plumbaginaceae, Persimmonaceae (Ebenaceae) , Styracaceae, Stink bug, Symplocaceae, Ash (Oleaceae), Loganiaceae, Gentianaceae, Menyanthaceae, Oleaceae, Apocynaceae ), Asclepiadaceae, Rubisaceae, Polemoniaceae, Convolvulaceae, Boraginaceae, Verbenaceae, Labiatae, Solanaceae, Scrophulariaceae ), Bignoniaceae, Acanthaceae, Sesame (Pedaliaceae), Fructose (Orobanchaceae). Gesneriaceae, Lentibulariaceae, Phrymaceae, Plantaginaceae, Caprifoliaceae, (Perox Adoxaceae), Valerianaceae, Dipsacaceae, Campanaceae ( Campanulaceae, Compositae, Myraceae, Juglandaceae, Salicaceae, Birchaceae, Beechaceae, Fagaceae, Ulmaceae, Mulberry, Nettle Urticaceae, Santalaceae, Mistletoe, Lothanthaceae, Polygonaceae, Apoaceae, Phytolaccaceae, Nyctaginaceae, Aizoaceae, Portulacaceae ), Caryophyllaceae, Chenopodiaceae, Amaranthaceae, Cactaceae, Magnoliaceae, Illiciaceae, Lauraceae, Cecidiphyllaceae, Minaria (Ranuncul aceae, Berberidaceae, Lardizabalaceae, Bird breeze (Amandiaceae, Menispermaceae), Nymphaeaceae, Ceratophyllaceae, Cabombaceae, Saururaceae , Piperaceae, Chloranthaceae, Aristolochiaceae, Actinidiaceae, Camellia, Theaceae, Guttaiferae, Droseraceae, Poppyaceae ), Capparidaceae, Cruciferaceae (Mustaceae, Cruciferae), Planeaceae (Plataceae, Platanaceae), Verruaceae, Hamamelidaceae, Pheasant (Snaphaceae, Crassulaceae), Panaxaceae (Saxifragaceae) Eucommiaceae, Pittosporaceae, Rosaceae, Leguminosae, Oxalidaceae, Geraniaceae, Tropaeolaceae, Zygophyllaceae, Linaceae Euphorbiaceae), star Callitrichaceae, Rutaceae, Simaroubaceae, Meliaceae, Polygalaceae, Anacardiaceae, Mapleaceae, Aceraceae, Mapleaceae, Mapleaceae (Hippocastanaceae), Sabiaceae, Balsam (Azalea, Balsaminaceae), Aquifoliaceae, Nova (Celastraceae), Staphyleaceae, Buxaceae, Empetraceae , Rhamnaceae, Vitaceae, Elaeocarpaceae, Tiliaceae, Malvaceae, Sterculiaceae, Adenaceae, Thymelaeaceae, Ellaeagnaceae, Flacourtiaceae, Violaceae, Passifloraceae, Tamaricaceae, Elatinaceae, Begoniaceae, Cucurbitaceae, Liliumaceae, Lythraceae Pomegranate Punicaceae, Onnagraceae, Haloragaceae, Bataceae (Alangiaceae), Dogwood (Hortisaceae, Cornaceae), Arboraceae (Araliaceae), Umbelaceae (Umbelliferae (Apiaceae) )), But is not limited thereto.

Most preferably the plant cells are selected from plants belonging to the Cruciferaceae, Rosaceae and Rosaceae.

The step of introducing the expression vector of step 2) into the host plant cell may be introduced into the plant by a known plant transformation method. That is, a well-known method known to those skilled in the art, such as Agrobacterium mediated method, gene gun, method using PEG, and electroporation, can be used. Preferably, the dicotyledonous plants are agrobacterium-mediated method (RB Horsch, et al., A simple and general method for transferring genes into plants, Science 227 (1985), pp. 1229-1231), Generation of DREB Transgenic Plants Using a Gene Gun (P. Christou, Particle Bombardment, Methods Cell Biol 50 (1995), pp. 375-382) or Agrobacterium Mediated (Oh et al. Plant Physiol 138: 341- 351, 2005; Ito et al. Plant Cell Physiol 47: 141-153, 2006).

After transformation, plant cells should be regenerated into whole plants. Techniques for the regeneration of mature plants from callus or protoplast cultures are well known for many different species (Handbook of Plant Cell Culture, Vol. 1-5, 1983-1989 Momillan, N.Y.). Thus, once transformation is realized, it is within the knowledge of the art to regenerate mature plants from transformed plant cells.

The foreign protein in the method of the present invention includes a protein that confers stress to the protein or transformant exerting a pharmacological effect.

As described above, since the swDREB1 gene derived from dried sweet potato silt of the present invention is strongly induced in various roots under various stress conditions including dryness, it is a trait that is resistant to environmental stress suitable for a disadvantageous condition. It can be usefully used to develop the converter.

Hereinafter, the present invention will be described in detail by way of examples.

However, the following examples are merely to illustrate the invention, but the content of the present invention is not limited to the following examples.

Example 1 Sweet Potato swDREB1  Gene cloning, sequencing and softness analysis

Ipomoea batatas (L.) Lam. Cv.White star plants were dried for 10 hours, and then the fibrous root tissues at 0, 0.5, 2, 6 and 10 hours were CTAB method (Kim and Hamada, Total RNA was isolated via Biotech Lett 27, 1841-1845, 2005). MRNA was isolated using a Poly (A) Tract mRNA separation system (Promega), and a cDNA library of dried sweet potato silt was prepared using SMART-cDNA synthesis kit (Clontech).

Five DREB genes were selected from the Arabidopsis Web site (http://www.arabidopsis.org) database for the production of DREB probe DNA for screening of DREB genes. Is the base sequence AtDREB1A / CBF3 : AT4G25480 (SEQ ID NO: 3: forward: 5'-ATGAACTCATTTTCTGCT-3 ', SEQ ID NO: 4: reverse: 5'-TTAATAACTCCATAACGATAC-3'), AtDREB1B / CBF1 : AT4G25490 (SEQ ID NO: 5: forward: 5'-TTTGGCTCCGATTAC-3 ', SEQ ID NO: 6 Reverse: 5'-TTAGTAACTCCAAAGCG-3'), AtDREB1C / CBF2 : AT4G25470 (SEQ ID NO: 7: Forward: 5'-ATGAACTCATTTTCTGCC-3 ', SEQ ID NO: 8 Reverse: 5 '-TTAATAGCTCCATAAGGAC-3'), AtDREB2A : AT5G05410 (SEQ ID NO: 9: Forward: 5'-ATGGCAGTTTATGATCAG-3 ', SEQ ID NO: 10: Reverse: 5'-TTAGTTCTCCAGATCCAA-3'), At DREB2B : AT3G11020 (SEQ ID NO: 11: Forward: 5'-ATGGCTGTATATGAACAAA-3 ', SEQ ID NO: 12 Reverse 5'-TCAAATATCCAGAGAACTC-3'). The library was screened by the conventional method using the prepared probe DNA, and 57 clones were obtained. DNA was isolated from the clone to determine the nucleotide sequence of the 5 'end of the cDNA using the T3 primer and obtain relevant genetic information and data through analysis of the Blast database of NCBI (www.ncbi.nlm.nih.gov) It was. Among them, DREB genes belonging to the DREB1 family of Arabidopsis were cloned as candidates for root-specific expression of various stresses, including drying, and the entire sequence of cDNA was determined. This cDNA gene was named ' swDREB1 ' cDNA.

The total length of the SwDREB1 cDNA gene is 1206 bp, which is composed of 132 bp 5 'terminal untranslated region (UTR), 300 bp 3' terminal untranslated region and 774 bp coding region (consisting of 257 amino acids). (FIG. 1). The NLS (nuclear localization signal) is a sequence for signaling to the nucleus of the DREB transcription factor, the AP2 / EREBP site is a DNA binding site, and the acidic region at the C terminus is a part that activates the DREB transcription factor. Compared with other DREB protein structures, the gene had well preserved NLS, AP2 / EREBP sites and C-terminal acidification sites. In particular, in the swDREB1 gene of the present invention, DSAWRL and LWSF, which are specific sequences of the DREB1 group, were preserved among the DREB family of Arabidopsis.

When the coding part of sweet potato swDREB1 was irradiated with BlastX, Capsicum showed CaCBF1B, the homology of 67% in CaDREBLP1 the amino acid level annuum), tomato (showed a homology of 61% than AtDREB1D / CBF4 of LeCBF1, Arabidopsis (Arabidopsis thaliana) in Lycopersicon esculentum) (Figure 2a). The relationship between the amino acid sequence of the putative protein of sweet potato swDREB1 and the DREB of various plant species was investigated using the ClustalW program (http://plant.pdrc.re.kr/gene/align/ClustalW.html). Is assumed to be a protein of the DREB1 family of Arabidopsis (FIGS. 2A and 2B).

Example 2 Sweet Potato Organization swDREB1  Gene expression analysis

RT-PCR was performed to analyze the tissue-specific expression patterns of swDREB1 , a sweet potato DREB transcription factor of the present invention, isolated from hay-treated sweet potato roots. Extract total RNA from sweet potato tissues (leaves, stems, storage roots, roots, coarse roots) using CTAB method (Kim and Hamada, Biotech Lett 27, 1841-1845, 2005), and then use 2.5 μg RNA to MMLV Reverse Transcription System cDNA. CDNA was synthesized using a synthetic kit (Clontech). SwDREB1 gene specific primers and the Accel Taq Premix kit (Genedocs) were used to investigate the expression of swDREB1 gene. As a specific primer of SwDREB1 , a nucleotide sequence (SEQ ID NO: 13: forward: 5'-CTATTCGCCCTATTCCTAT-3 ', SEQ ID NO: 14: reverse: 5'-CAAATTCAAGCCTTGGTATC-3') was used.

As a result, the swDREB1 gene was strongly expressed in the tuberous root and stem, and also in the fibrous root. However, the leaf and the thick pigmented root (thick pigmented root) showed a very weak appearance (Fig. 3). Thus, swDREB1 of the present invention isolated from dried sweet potato root tissue The gene was found to be a gene expressed in stems, roots, and storage roots.

<Example 3> by various stress treatment including drying treatment swDREB1  Gene expression analysis

In order to analyze the degree of response of the swDREB1 gene to various stress treatments including drying, the expression changes of swDREB1 gene were analyzed after various stress treatments such as drying, plant hormone (ABA), temperature and chemicals. Analyzed by PCR.

First, swDREB1 In order to analyze the degree of gene response by the drying treatment, the leaves and the roots of sweet potatoes were dried by time zones (0, 1, 2, 8, 16, 24 hours) and then the method was performed in Example 2 After RNA was isolated, cDNA was synthesized, respectively. SwDREB1 used in Example 2 RT-PCR was performed with gene specific primers. As a result, the swDREB1 gene showed the strongest expression level at 1 hour after drying in the root root tissue, and maintained strong expression level until 16 hours (FIG. 4A). On the other hand, no increase or decrease in the expression level of swDREB1 was observed in the leaf tissues.

In general, under dry stress, the amount of plant hormone abscis acid is increased, and it is known that abscis acid controls pore opening and closing of plants in response to drying. In addition, DREB transcription factors of various plant species are classified as DREB transcription factors with abscis acid dependent or independent signaling mechanisms, depending on whether the expression is responsive by treatment with abscis acid. Therefore, to observe whether the swDREB1 gene in response to drying treatment is increased by the application of absic acid, the sweet potato leaves and silt tissues were treated with 0.1 mM abscis acid at time intervals (0, 12, 24, 36, 48 hours). After treatment, RNA was extracted and cDNA was synthesized in the same manner and RT-PCR was performed. As a result, no increase or decrease in the amount of swDREB1 expression by abscitic acid treatment was observed in the leaf and silt tissue (FIG. 4B). Thus, the sweet potato swDREB1 gene is thought to be regulated by signaling mechanisms independent of absic acid.

Arabidopsis DREB genes showed low temperature and salt induced response characteristics. In order to see the responsiveness of swDREB1 to temperature stress, various low-temperature stresses were treated at 15 ° C. and 4 ° C. for 16 hours in an incubator condition, after which RNA was isolated and cDNA was synthesized and RT-PCR was performed. As a result, the swDREB1 gene showed the strongest expression level after low temperature (4 ° C.) treatment in leaf and root root tissues (FIG. 4C).

Next, 100 mM sodium chloride to see the reactivity to various compounds stress of swDREB1 and 0.05 mM methyl rain halogen (methyl viologen, paraquat) 24 hours after, and 0.05 mM Cd 48 hours post-treatment post-treatment to extract RNA by the same method, and cDNA was synthesized and RT-PCR was performed. Methyl viologen is a non-selective herbicide that generates excess of free radicals to kill plants. Sterile water was used for each control. As a result swDREB1 Genes showed strong expression increase in sodium chloride, methyl viologen, cadmium treatment in leaf and silt tissue (FIG. 4D). Therefore, it can be seen that the swDREB1 gene has responsiveness to various stresses, and in particular, high expression level in the roots. Therefore, the swDREB1 gene is expected to help the development of environmentally resistant plants (including sweet potatoes) that improve early growth of seedlings in poor soil such as dry and cold.

Example 4 S wDREB1  Southern blot analysis of genes

Southern blot analysis was performed to observe the swDREB1 gene of the present invention in the sweet potato genome and the presence of other homologous genes. Genomic DNA was isolated and purified from sweet potato leaves by CTAB method, and then digested with Eco RI, Eco RV, and Hind III. After electrophoresis of genomic DNA digested with restriction enzymes, a Southern blot was performed using a probe labeled 300 bp of the 3 'terminal specific region of swDREB1 cDNA with a ³² P radioisotope.

As a result swDREB1 gene It was found that the swDREB1 gene of the present invention to DNA bands ride chimja recognizes the swDREB1 exists one in each fragment cut with restriction enzymes that are present in a single gene in the potato genome (Fig. 5 ).

1 is a view showing the nucleotide sequence of the dried sweet potato root derived swDREB1 gene of the present invention and the amino acid sequence deduced therefrom.

Figure 2a is a view comparing the amino acid sequence of the deduced protein of the swDREB1 gene of the present invention and the DREB gene amino acid sequence of various plants (pepper, tomato, Arabesque , cotton, sweet cherry)

Figure 2b is a view showing the flexible relationship between the amino acid sequence of the deduced protein of the swDREB1 gene of the present invention and the DREB gene of various plants (pepper, tomato, Arabidopsis, cotton, sweet cherry).

Figure 3 is a photograph of RT-PCR electrophoresis of the expression of the swDREB1 gene of the present invention in various tissues of sweet potatoes. L, leaf; S, stem; TR, tuberous root (storage root); FR, fibrous root (root); TPR, thick pigmented root.

Figure 4a is a picture of RT-PCR electrophoresis showing the expression pattern of the swDREB1 gene of the present invention at 0, 1, 2, 8, 16, 24 hours after drying the sweet potato leaves and roots.

Figure 4b is a RT-PCR electrophoresis picture showing the expression pattern of the swDREB1 gene of the present invention at 0, 6, 12, 24, 48 hours after treatment with 0.1 mM of abscitic acid in sweet potato leaves and sils roots .

Figure 4c is a picture of RT-PCR electrophoresis showing the expression pattern of the swDREB1 gene of the present invention 16 hours after the sweet potato leaves and the roots were subjected to temperature stress (4 ℃, 15 ℃).

FIG. 4D shows the present invention at 24 hours after treatment with 100 mM sodium chloride (NaCl) and 0.05 mM methyl viologen on sweet potato leaves and roots, and 48 hours after treatment with 0.5 mM cadmium. RT-PCR electrophoresis picture of the swDREB1 gene expression.

5 is a Southern blot photograph confirming that the swDREB1 gene of the present invention exists on the genome of sweet potatoes.

<110> Korea Research Institute of Bioscience and Biotechnology <120> SwDREB1 protein originated from a root of Ipomoea batatas and          gene coding the same <130> 7p-11-07 <160> 14 <170> KopatentIn 1.71 <210> 1 <211> 1206 <212> DNA <213> SwDREB1 gene from a root of Ipomoea batatas <400> 1 aatacatacc cacttccagt tttagctttt ccatatatat atatatatat actttactaa 60 gtaccgccgt cgtttacgta cctactacta gtctcaatca ctcaagattc gattctatat 120 atggatatgg atatggatat agttgggaat tattattctg ggaattttct aagtgctgct 180 gcggcggcgt cgagtttttg gtcgccggaa atgggtgcag tagtgccttc gccactgtct 240 tcttctgata ctgggagttg cagcgctact atgaaagcga atttgtcgga tgaagaggtg 300 ttgttggctt ctaataatcc gaagaaacgc gctgggagga agaagtttcg ggagactcga 360 cacccggtgt accggggagt gaggaggagg aactccggga agtgggtgtg tgaggtgagg 420 gagcccaaca agaagtccag gatatggctg ggaactttcc ccacggctga aatggcggct 480 agagctcatg acgtggccgc catcgctctc agaggctgct ccgcctgtct caacttcgcc 540 gactcggctt ggaggcttcc aatcccggcg tccgccgacc ccaaggacat ccagaaagct 600 gcggcggagg cggcggaggc tttccgtcca gtggcactgc cagcaaacca aaaccaaacc 660 caacggatta ttctagaagc tgaagaagaa gaagaagagt gcaacagtag tatgaaagag 720 gaacaagtgt ccacaaccaa cgaaaacgtg ttcttcatgg acgaggaagc gtttttcgac 780 atgcccggat tgcttgctga catggctcaa gccctgatgc tacctccacc tcaatgcgca 840 ctagtggacc gtaccaatga cgtggagctt gatgctgacg tgtcactctg gtctttctcc 900 atttaaaacc caaaattagg atgaagatgg ataatctatc tgtgaattga atgtcgtcca 960 ttgtccactc tactctactc cgtacgtact agtctctagt tatatgaata gaagggatgc 1020 ctttcaagaa aaggcgaaaa gcgtccattc ctattcgccc tattcctatt cctattctat 1080 ataggtagga ttagctaata tccgtacttc attttttgag gttgtacggc ttgtaaaata 1140 tccgatacca aggcttgaat ttgaatgaat tatgccaaaa aaaaaaaaaa aaaaaaaaaa 1200 aaaaaa 1206 <210> 2 <211> 257 <212> PRT <213> SwDREB1 protein from a root of Ipomoea batatas <400> 2 Met Asp Ile Val Gly Asn Tyr Tyr Ser Gly Asn Phe Leu Ser Ala Ala   1 5 10 15 Ala Ala Ala Ser Ser Phe Trp Ser Pro Glu Met Gly Ala Val Val Pro              20 25 30 Ser Pro Leu Ser Ser Ser Asp Thr Gly Ser Cys Ser Ala Thr Met Lys          35 40 45 Ala Asn Leu Ser Asp Glu Glu Val Leu Leu Ala Ser Asn Asn Pro Lys      50 55 60 Lys Arg Ala Gly Arg Lys Lys Phe Arg Glu Thr Arg His Pro Val Tyr  65 70 75 80 Arg Gly Val Arg Arg Arg Arg Asn Ser Gly Lys Trp Val Cys Glu Val Arg                  85 90 95 Glu Pro Asn Lys Lys Ser Arg Ile Trp Leu Gly Thr Phe Pro Thr Ala             100 105 110 Glu Met Ala Ala Arg Ala His Asp Val Ala Ala Ile Ala Leu Arg Gly         115 120 125 Cys Ser Ala Cys Leu Asn Phe Ala Asp Ser Ala Trp Arg Leu Pro Ile     130 135 140 Pro Ala Ser Ala Asp Pro Lys Asp Ile Gln Lys Ala Ala Ala Glu Ala 145 150 155 160 Ala Glu Ala Phe Arg Pro Val Ala Leu Pro Ala Asn Gln Asn Gln Thr                 165 170 175 Gln Arg Ile Ile Leu Glu Ala Glu Glu Glu Glu Glu Glu Cys Asn Ser             180 185 190 Ser Met Lys Glu Glu Gln Val Ser Thr Thr Asn Glu Asn Val Phe Phe         195 200 205 Met Asp Glu Glu Ala Phe Phe Asp Met Pro Gly Leu Leu Ala Asp Met     210 215 220 Ala Gln Ala Leu Met Leu Pro Pro Pro Gln Cys Ala Leu Val Asp Arg 225 230 235 240 Thr Asn Asp Val Glu Leu Asp Ala Asp Val Ser Leu Trp Ser Phe Ser                 245 250 255 Ile     <210> 3 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> AtDREB1A / CBF3: AT4G25480 forward primer <400> 3 atgaactcat tttctgct 18 <210> 4 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> AtDREB1A / CBF3: AT4G25480 reverse primer <400> 4 ttaataactc cataacgata c 21 <210> 5 <211> 15 <212> DNA <213> Artificial Sequence <220> <223> AtDREB1B / CBF1: AT4G25490 forward primer <400> 5 tttggctccg attac 15 <210> 6 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> AtDREB1B / CBF1: AT4G25490 reverse primer <400> 6 ttagtaactc caaagcg 17 <210> 7 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> AtDREB1C / CBF2: AT4G25470 forward primer <400> 7 atgaactcat tttctgcc 18 <210> 8 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> AtDREB1C / CBF2: AT4G25470 reverse primer <400> 8 ttaatagctc cataaggac 19 <210> 9 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> AtDREB2A: AT5G05410 forward primer <400> 9 atggcagttt atgatcag 18 <210> 10 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> AtDREB2A: AT5G05410 reverse primer <400> 10 ttagttctcc agatccaa 18 <210> 11 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> AtDREB2B: AT3G11020 forward primer <400> 11 atggctgtat atgaacaaa 19 <210> 12 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> AtDREB2B: AT3G11020 reverse primer <400> 12 tcaaatatcc agagaactc 19 <210> 13 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Forward primer specific for swDREB1 <400> 13 ctattcgccc tattcctat 19 <210> 14 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Reverse primer specific for swDREB1 <400> 14 caaattcaag ccttggtatc 20  

Claims (14)

Sweet potato derived from having an amino acid sequence set forth in SEQ ID NO: 2 swDREB1 protein. A polynucleotide encoding the swDREB1 protein of claim 1. The polynucleotide according to claim 2, which has a nucleotide sequence set forth in SEQ ID NO: 1 . An expression vector comprising the polynucleotide of claim 2. A transformed host cell transformed with the expression vector of claim 4. 1) preparing a transformed plant cell by introducing the expression vector of claim 4 into a host plant cell; 2) culturing the prepared transgenic plant cells to prepare a callus (callus); And 3) A method of producing a transformed plant having a stress resistance, comprising the step of inducing differentiation of the prepared callus. The method of claim 6, wherein the stress is induced by drying, abscidic acid, low temperature, sodium chloride, methyl viologen or cadmium. delete The method of claim 6, wherein the plant cell is selected from the group consisting of plants belonging to a cruciferaceae, an eggplant, a rose family, and an asteraceae. A plant which is a progeny or clone of a transgenic plant produced by the method of claim 6. Callus of a transgenic plant produced by the method of claim 6. 1) preparing a recombinant expression vector comprising a polynucleotide as set forth in SEQ ID NO: 1 and a polynucleotide encoding a foreign protein operably linked thereto; 2) preparing a transgenic plant cell by introducing the recombinant expression vector into a host plant cell; 3) culturing the prepared transgenic plant cells to prepare a callus; 4) preparing a transformed plant by inducing differentiation of the prepared callus; And 5) A method for high expression of a foreign protein in the root, comprising the step of culturing the transformed plant prepared. The method of claim 12, wherein the plant cells are selected from the group consisting of plants belonging to the cruciferaceae, the branch family, the Rosaceae family, and the Conifer family. The method of claim 12, wherein the foreign protein is a protein expressing resistance to stress to a protein or transformant exerting a pharmacological effect.

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