KR20100038627A - Pepper disease resistance- and cell death-related, abscisic acid-responsive gene caabr1 and transgenic plants using the same - Google Patents

Abstract

PURPOSE: A CaABR1(Capsicum annuum Abscisic acid-responsive protein 1) relating plant disease resistance and apoptosis and to produce resistant plant and transgenic plant. CONSTITUTION: A Capsicum annuum L. cv. Nockwang-derived gene, CaABR1 has a sequence of sequence number 1. An isolated protein CaABR1 has an amino acid of sequence number 2. A recombinant vector pBIN35S::CaABR1 is prepared by inserting CaABR1 gene into pBIN35S. A transgenic plant is obtained by transforming CaABR1 gene with the recombinant vector. The plant is Capsicum annuum or the arabidopsis thaliana. The resistance of the plant is analyzed by isolating RNA from a plant which is inoculated with non-pathogenic bacteria and confirming CaABR1 gene expression by Northern blot.

Description

Pepper disease resistance- and cell death-related, abscisic acid-responsive gene CaABR1 and transgenic plants using the same}

The present invention relates to a plant disease resistance and apoptosis-related Epsic acid response gene CaABR1 ( Capsicum annuum Abscisic acid-responsive protein 1 ) isolated from red pepper ( Capsicum annuum L.) plants, and a method for screening disease resistance in plants using the same The invention relates to the production of converted plants. More specifically, the present invention provides a nucleotide sequence and amino acid sequence of CaABR1 gene, a novel gene associated with plant disease resistance and apoptosis, isolated from pepper greenery varieties after infection with pathogens. It provides a resistance search method for searching for differential expression of genes. In addition, the present invention provides a method for temporarily overexpressing this gene in a red pepper plant, a virus-induced gene, via a recombinant vector (pBIN35S: CaABR1) using the CaABR1 gene and Agrobacterium- mediated transient expression. This gene is not expressed through virus-induced gene silencing (VIGS), and is introduced and overexpressed in Arabidopsis thaliana plants to enhance plant disease resistance of CaABR1 gene. The present invention relates to providing a transformed plant through this.

Red pepper is one of the most used vegetables in Korea. However, in the environment-friendly pepper cultivation, a problem has been raised in the stable supply and demand of peppers due to the reduction of pepper production and quality loss caused by the occurrence of pepper disease. Although the use of chemical pesticides in general pepper cultivation is becoming common to suppress the occurrence of pepper disease, the problem of human residue, environmental toxicity, and pollution of pesticides is occurring. The pepper cultivation technology is developing in the direction of bottle control.

     At present, the most preferred method of controlling pepper disease is breeding and growing disease-resistant varieties. In order to cultivate such disease-resistant pepper varieties, it would be efficient to breed pepper varieties by molecular breeding to find useful pepper-related genes.

On the other hand, plants are known to have physical barriers such as strengthening cell walls externally and forming callus or killing themselves to prevent further proliferation in order to prevent (resist) pathogen invasion.

In addition, changes in hormones in the plant occur and various reactions occur according to the changes in the hormones. Abscisic acid, a type of plant hormone, has been reported to cause resistance reactions depending on the type of pathogen and plant when internally increased or treated externally. In plants, there are numerous genes induced by this hormone, each of which has different expressions in different organs and is known to be specifically induced against biological and abiotic environmental stresses.

By identifying the functions of Epsic acid-responsive genes, which are not yet known in pepper plants, this study can be used to study the defense mechanism against plant diseases in pepper plants. These genes are useful for the molecular breeding of pepper disease resistant varieties. It can be used as a resource.

The present invention is to solve such a prior art problem, the purpose of which is involved in the defensive reaction against bacterial bacterial spot disease ( Xanthomonas campestris pv.vesicatoria ), one of the genes involved in plant disease resistance and cell death Epsil By separating and deciphering the acidic acid-responsive protein gene 1 ( CaBR 1 ), the nucleotide sequence information and amino acid sequence information thereof are provided.

In addition, the present invention is to provide a resistance search method for searching for the presence of resistance to plant diseases and the change of plant hormones using the CaABR1 gene. Furthermore, the aim is to introduce a transgenic plant having resistance to plant diseases by introducing the gene into the plant.

Other objects and advantages of the present invention will become apparent from the following detailed description, claims and drawings.

The above object of the present invention, inoculated with pepper bacterium bacillus bacterium ( Xanthomonas campestris pv.vesicatoria) in the capsicum annuum cv.Nockwang , new resistance and cell death-related Epsic acid reaction isolated from the inoculated pepper leaves The gene CaABR1 was identified and its nucleotide sequence was determined. Using this, CaABR1 gene was examined by pathogen inoculation and epsilic acid treatment, and CaABR1 gene in Arabidopsis thaliana overexpressed red pepper plants and this gene. Achieved by confirming the plant disease resistance induction function of.

Accordingly, the present invention provides the amino acid sequences of the isolated gene CaABR1 and SEQ ID NO: 2 having the nucleotide sequence of SEQ ID NO: 1 related to plant disease resistance and apoptosis-related Epsic acid reactions derived from Capsicum annuum L. cv. Nockwang. To provide an isolated epsic acid reactive protein (CaABR1).

In another aspect, the present invention provides a recombinant vector and a transformed plant prepared using the gene.

Furthermore, the present invention provides a method for searching for disease resistance or plant hormone changes in plants, including the steps of identifying RNAs from plants inoculated or treated with pathogens or epsis acid and confirming differential expression of CaABR1 gene by Northern blot analysis. .

Hereinafter, with reference to the accompanying drawings will be described in detail the configuration of the present invention.

First, the present invention is to identify proteins that are differentially expressed by extracting the protein from the cultivars of the green No. AD which exhibits hypersensitivity resistance to the inoculation of non-pathogenic strains of red pepper bacterial bacillus bacteria ( Xanthomonas campestris pv.vesicatoria ). After determining the complete gene sequence from the cDNA library prepared by separating the mRNA from the inoculated leaves using the degree of the identified partial sequence degree, the epsilic acid-responsive protein coding gene sequence (SEQ ID NO: 1) of Caybean greenery varieties named CaABR1 and According to amino acid sequence (SEQ ID NO: 2).

CaABR1 protein according to the present invention was found to have a single GRAM domain that appears in several eukaryotes as a result of the analysis of the amino acid sequence, and the nucleotide with the unnamed protein product gene (Accession no. CAO41288) of the vine ( Vitis vinifera ) (74%) at the nucleotide level and 63% homology with the GRAM domain-containing protein gene (Accession no. NP_196824) of Arabidopsis thaliana . http://blast.ncbi.nlm.nih.gov/Blast.cgi, nucleotide blast program). The gene sequence of the identified CaABR1 and the amino acid sequence encoded by this sequence are shown in FIG.

Next, the present invention provides a method for detecting resistance of plants consisting of confirming whether CaABR1 protein is expressed by applying various organs, pathogens and hormonal induction methods of pepper plants, and performing RNA Northern gel blots. More specifically, the present invention relates to a method for detecting resistance of plants, comprising checking whether CaABR1 protein is expressed by using the CaABR1 protein gene as a probe for a selected organ in a pepper individual inoculated or treated with a pathogen or epsis acid. to provide. It was confirmed through experiments that the CaABR1 gene according to the present invention is differentially induced and expressed by phytopathogens or epsic acid (see FIGS. 3 and 4). Therefore, the CaABR1 gene according to the present invention can be usefully used for a method of searching for resistance of plants.

In addition, the present invention by using the recombinant vector containing the CaABR1 gene, the production method of new disease-resistant transformed plants and resistance molecular breeding by confirming whether the plant disease resistance generation and cell death of pepper plants and transgenic Arabidopsis plants It can provide the material of. More specifically, the present invention uses a recombinant vector (pBIN35S :: CaABR1 ) containing the CaABR1 gene (FIG. 5) and the Agrobacterium tumefaciens strain GV3101 strain, transient expression in pepper plants. ) method, VIGS plant production and Arabidopsis for inhibiting the expression of CaABR1 gene (Arabidopsis thaliana), transgenic expression in plants (transgenic expression) to check the plant disease resistance activation function of CaABR1 gene through (see Fig. 6a to Fig. 15 ), Thereby providing a transgenic plant.

As described above, by completing the present invention, CaABR1 encoding Epsic acid reactive protein, which is one of genes related to plant disease resistance, is involved in the defense against pepper plant bacterial disease ( Xanthomonas campestris pv.vesicatoria ). By separating and deciphering the gene, the sequence information and amino acid sequence information are provided, as well as providing a method for detecting plant disease resistance and hormonal response using this gene, and contributing to the production of resistant plant breeding and transgenic plants. It became possible to plan the effect.

In addition, by overexpressing the CaABR1 gene according to the present invention in pepper and Arabidopsis to identify plant disease resistance and conversely, to identify the resistance reduction through CaABR1 gene silencing provides a method for detecting plant disease resistance and CaABR1 according to the present invention It can be provided as a material for breeding resistant plants.

The present invention enables the accumulation of genetic information of the response to the disease and related disease-associated protein (PR protein) can provide an interesting model for signaling of the disease in plants, biochemical changes that cause resistance By understanding this, it may be practically used to develop genetically engineered plants that promote disease resistance or to develop plant protection chemicals that operate to promote the plant's genetic resistance. Therefore, the present invention has an effect that can promote the development of resistant varieties is a very useful invention in the plant breeding industry.

Hereinafter, the present invention will be described in more detail with reference to the accompanying drawings. These examples are only for illustrating the present invention in more detail, it will be apparent to those skilled in the art that the scope of the present invention is not limited by these examples in accordance with the gist of the present invention. .

Example 1. CaABR1 Gene base sequence and amino acid sequence determination

Pepper disease resistance-related response of epsi seusan one protein for bacterial jeommunuibyeong protein (CaABR1: C apsicum a nnuum Ab scisic acid- r esponsive 1 protein) coding gene sequences and tested as described below to provide the amino acid sequence deduced therefrom Was performed.

Nokgwang pepper cultivars (Capsicum annuum cv. Nockwang) jeommunuibyeong pepper bacterial non-pathogenic bacteria strain (Xanthomonas campestris pv. Vesicatoria Bv5-4a) for inoculation, and then brought out of the treatment appeared seupsil seupsil resistant lesions of apoptosis sensitization reaction for 18 hours, the plants Leaves were harvested from healthy plants without inoculation with.

MRNA was extracted from the collected leaves and reverse transcribed, followed by PCR (Polymerase Chain Reaction) amplification to prepare cDNA libraries. In addition, proteins were extracted from the collected leaves and subjected to two-dimensional gel electrophoresis.

As a result of two-dimensional gel electrophoresis, the protein specifically increased in only the protein gel of the plant inoculated with the non-pathogenic strain was assumed to be a protein related to pathogenic resistance. Identification was performed using mass spectrometry.

Gene sequences cloned into GenBank using the Blast program after cloning and sequencing complete genes from cDNA libraries using known mRNA sequence information (Accession no. CA524559) of the identified proteins. Compared with the CaABR1 protein coding gene was identified and named as epsis acid response gene 1 ( CaABR1 ).

The CaABR1 protein according to the present invention was found to have one GRAM domain in several eukaryotes as a result of analysis of the amino acid sequence, and with the unnamed protein product gene (Accession no. CAO41288) of Vitis vinifera It was found to be novel by showing a homology of 74% at the nucleotide level and 63% with the GRAM domain-containing protein gene (Accession no. NP_196824) of Arabidopsis thaliana ( http://blast.ncbi.nlm.nih.gov/Blast.cgi, nucleotide blast program). The confirmed gene base sequence of CaABR1 and the amino acid sequence encoded by this sequence are shown in FIG.

Example 2 Peppers in Various Pepper Organs CaABR1  Exploration of Gene Expression

In order to confirm the expression of CaABR1 gene obtained in Example 1 in the pepper organ was carried out the following test.

RNA was isolated from each organ of pepper, leaves, stems, roots, flowers, green fruits, red fruits, and the like. The isolated RNA was electrophoresed on a formaldehyde-added gel and then transferred to a nylon membrane (Hybond N +).

The CaABR1 gene obtained in Example 1 was labeled with ³² P-dCTP and then this was reacted for 18 hours at 65 ° C. in a reaction solution [0.25 M phosphate buffer, 7% SDS, 1 mM EDTA, 5% dextran sulfate]. Culture was performed to hybridization (hybridization).

The ³² P-labeled DNA probe was attached to the mRNA of the nylon membrane. When the X-ray film was placed on the membrane, the ^32- P labeled DNA was exposed to the X-ray film to detect the expression of the gene. .

It was shown in Figure 2 whether the RNA Northern gel blot was loaded with the same amount of ribosomal RNA, the experimental results showed that the CaABR1 gene is expressed in the stem, green fruit, red fruit.

Example 3. By Plant Hormone Epsis Acid Treatment CaABR1  Confirmation of gene expression

In order to determine whether the CaABR1 gene of the present invention is expressed in response to epsic acid like other genes having homology, the following test was performed.

To 100 u M concentration of epsi seusan 6 treated using a leaf stage of the pepper sprayer to plant foliage, and collected by the blade 6, 12, 18, 24 hours after treatment to extract RNA and as in Example 2 using the RNA RNA gel blots were performed in the same manner.

As a result of the experiment, CaABR1 gene was shown to start expression after 6 hours as shown in FIG. 3 and remained strong at 12 hours and then decreased.

Example 4 In Pepper Plants by Pathogen Inoculation CaABR1  How to Explore Expression of Genes

In order to confirm the method of using the CaABR1 gene obtained in Example 1 for the resistance search was tested as follows.

First, in order to present a specific resistance screening method, after incubating the non-pathogenic strain Bv5-4a with an incompatible reaction with the pathogenic strain Ds1 of the pepper bacterial bacillus bacterium ( Xanthomonas campestris pv.vesicatoria ), which has a friendly reaction with the plant, 10 ⁸ cfu / Ml of red pepper varieties 3 and 4 leaves of the red pepper varieties at the concentration of 6 ml in the infiltration using a syringe (infiltration), and the wet treatment for 18 hours at 28 ℃ temperature and 100% relative humidity conditions. After inoculation, 1, 5, 10, 15, 20 and 25 hours after inoculation, 3 and 4 leaves were inoculated respectively to separate RNA and RNA gel blot was performed as in Example 2.

It is shown in Figure 4 that the ribosomal RNA amount is equally loaded when performing the RNA gel blot, and the results show that the CaABR1 gene is expressed when inoculated into a non-pathogenic strain while the resistance reaction occurs up to 20 hours after inoculation. This seemed to increase strongly and decrease.

Example 5. CaABR1  Changes in Pepper Plants due to Temporary Overexpression of Genes

35S: CaABR1 was prepared by introducing the CaABR1 gene obtained in Example 1 into the pBIN35S vector provided by Stratagene to determine what changes occur when CaABR1 gene is overexpressed in pepper plants (FIG. 5).

The recombinant vector thus prepared was introduced into Agrobacterium tumefacien strain GV3101 through electroporation and inoculated onto pepper leaves to overexpress CaABR1 gene.

In the CaABR1 transient expression experiment, a control vector (35S: 00) without a gene was used as a control. GV3101 strain containing these two vectors was shaken in Yeast-Peptone (YEP: yeast extract 10 g, peptone 10 g, sodium chloride 5 g / L) medium and absorbed by the spectrophotometer. (Optical density) 600 (OD ₆₀₀ ) was adjusted to 0.5, 0.2, 0.1, 0.05 and inoculated at the border of the leaf veins of pepper, respectively, and the results are shown in Figures 6a and 6b.

As a result, cell death was observed in the region overexpressing the CaABR1 gene at 36 hours after inoculation (first photo of FIG. 6A), and trypan blue staining was performed to clearly observe cell death. As a result, it was observed that the sites where cell death occurred were blue stained (third photo of FIG. 6A). In addition, DAB (DA, 3,3'-Diaminobenzidine) staining was performed to investigate the change in free radicals due to overexpression of this gene. Agrobacterium (Agrobacterium) increase due to due to the vaccination experiment sphere and over-expression of only occur that the accumulation of reactive oxygen species in both CaABR1 control did not appear distinctly (middle picture of Figure 6a).

Conductivity was measured using an ion meter to specifically quantify cell death. The inoculated portion at the concentration of OD ₆₀₀ = 0.2 in the leaves inoculated with each experimental and control group was cut out in a circular shape, washed once in distilled water, and immersed in fresh distilled water again, and then the conductivity was measured after 30 minutes. As a result, the CaABR1 gene was overexpressed and the cell death occurred in the leaves where there was a lot of ion leakage, and thus the conductivity was higher than that of the control (Fig. 6b).

In order to observe the expression of disease resistance-related genes in CaABR1 overexpressed pepper leaves, the inoculated leaves were harvested and RNA was extracted and real time RT-PCR was performed and the results are shown in FIG. 7. . As shown in FIG. 7, the expression of CaBPR1 ( Capsicum annuum basic pathogenesis-related protein 1), CaDEF1 ( Capsicum annuum defensin 1), etc., reported as pathogenic genes, was increased at CaABR1 overexpression sites. .

In order to investigate whether callus formation known as a defense mechanism was increased in CaABR1 overexpressed red pepper leaves, the leaf region overexpressed with CaABR1 gene was stained with aniline blue. As shown in FIG. 8, it was observed that the callus also increased with the concentration of GV3101 at the CaABR1 overexpression site.

Example 6. Using Viruses CaABR1  Silencing of Genes and Changes in Disease Resistance

To investigate the changes in disease resistance when CaABR1 genes lost function in red pepper plants, the genes were silenced using TRV1 and TRV2 vectors derived from Tobacco rattle virus and tested as follows. Was performed.

First, the CaABR1 gene was introduced into the TRV2 vector to prepare a TRV: CaABR1 recombinant vector. More specifically, embodiments of CaABR1 gene obtained from one virus-induced gene silencing (virus-induced gene silencing, VIGS) TRV2 the CaABR1 gene using the TRV (Tobacco Rattle Virus) which is used in the vector (reference: Baulcombe DC (1999) Fast forward genetics based on virus-induced gene silencing.Curr. Opin.Plant Biol. 2: 109-113) to prepare a recombinant vector TRV2 :: CaABR1 .

Thus prepared TRV: CaABR1 recombinant vector was transformed into the GV3101 strain as in Example 5. A vector without this gene (TRV: 00) was transformed into the GV3101 strain for use as a control. It was also transformed with a TRV1 vector (Bulcombe DC (1999) Fast forward genetics based on virus-induced gene silencing.Curr. Opin. Plant Biol. 2: 109-113) to be used together.

Incubate the strains with these vectors as in Example 5, adjust the concentration from OD ₆₀₀ to 0.4, mix the strain with TRV1 and the strain with TRV: CaABR1 1: 1, and then use a syringe when the cotyledon of the pepper appeared. A plant was inoculated with cotyledon to silence the CaABR1 gene. For control, strains with TRV1 and strains with TRV: 00 were similarly inoculated.

When the red pepper thus prepared reached the six leaf stage , it was inoculated with red pepper bacterial spot bacteria ( Xanthomonas campestris pv. Vesicatoria ) to determine whether the disease resistance was reduced. 9A and 9B, when inoculated with the pathogenic strain (Ds1), it was observed that the symptom was large in the plants in which the CaABR1 gene was silenced, and the growth rate of the bacteria was increased. However, no change in bacterial growth rate by epsis acid was observed in plants treated with epsis acid one day before inoculation. In addition, when inoculated with a non-pathogenic strain (Bv5-4a), it was observed that the bacterial growth rate increased as the apoptotic response was delayed in the caABR1 gene- silent plants (FIGS. 10A and 10B). Epsis acid inhibited bacterial growth in plants without CaABR1 silencing, but treatment of epsis acid in plants with this gene silenced increased bacterial growth (FIG. 10B). Since CaABR1 delayed apoptosis in silenced plants, the conductivity was measured as in Example 5, indicating that the gene has lower conductivity than non- silenced plants in silenced plants, resulting in reduced cell death and reduced ion leakage. Observation was possible (FIG. 10C).

Example 7. CaABR1 Preparation of Transgenic Plants Using Genes CaABR1  Disease Resistance Changes in Arabidopsis Plants by Overexpression of Genes

In order to determine whether the disease resistance was increased when the CaABR1 gene was overexpressed, pepper CaABR1 gene was transferred to Arabidopsis plants that were easily transformed by the floral dipping method of the GV3101 strain having the recombinant vector used in Example 5. After transformation by transformation, the following test was performed.

In order to determine the change in disease resistance in plants overexpressing the CaABR1 gene, CaABR1 overexpression was first observed in the Arabidopsis larvae overexpressed with the CaABR1 gene and wild-type Arabidopsis as a control, and the results are shown in FIG. 11. # 1, # 2 and # 3 represent the line number of the transgenic plant used.

Pseudomonas siryngae pv. Tomato in the Arabidopsis plants overexpressed CaABR1 as described above DC3000 strains were inoculated using a syringe at a concentration of 5 x 10 ⁴ cfu / ml and placed at 28 ° C to observe changes in the plant. As shown in FIG. 12, it was observed that the growth rate of bacteria in the overexpressed plants of CaABR1 showed resistance as compared to the wild type.

In addition, Hyaloperonospora parasitica Noco2 strain, which causes downy mildew, was inoculated at 5 x 10 ⁴ spores / ml at the cauliflower season of CaABR1 overexpressing Arabidopsis plants and wild-type control Arabidopsis plants, and then wetted at 17 ° C. After treatment, changes in disease resistance were observed.

As a result, as shown in FIG. 13, less sporangiophore formation was observed in plants overexpressed CaABR1 gene than in wild-type control plants (FIG. 13). In addition, as a result of confirming spore hard water in the cotyledon of each plant, less spore hard water was formed in the transgenic Arabidopsis plants overexpressed the CaABR1 gene (FIG. 14). Furthermore, each plant cotyledon was collected, suspended in 1 ml of sterile water, and then strongly released to confirm the conidiospore number. As a result, the number of conidia was observed to be reduced in plants overexpressing the CaABR1 gene (FIG. 15).

Although specific portions of the present invention have been described in detail above, it will be apparent to those skilled in the art that such specific techniques are merely preferred embodiments, and thus the scope of the present invention is not limited thereto. Therefore, the substantial scope of the present invention will be defined by the appended claims and equivalents thereof.

1 is a view showing the nucleotide sequence and amino acid sequence of pepper Epsis acid reaction gene cloned from the pepper green light varieties ( Capsicum annuum L. cv. Nockwang) according to the present invention,

Figure 2 is a diagram showing the expression results of CaABR1 gene in each organ of the red pepper cultivation varieties,

Figure 3 is a view showing the expression results of CaABR1 gene over time after the treatment with Epsis acid (Abscisic acid) to pepper plants,

4 is a view showing the expression results of CaABR1 gene in the inoculated and non-inoculated leaves when inoculated with the pathogenic and non-pathogenic strains of pepper bacterial erythematophytes ( Xanthomonas campestris pv.vesicatoria ) to the red pepper varieties,

5 is an arrangement map of a recombinant vector (pBIN35S: CaABR1 ) for overexpressing the CaABR1 gene in pepper and Arabidopsis thaliana plants,

6a and 6b show the recombinant vector pBIN35S: CaABR1 introduced into the Agrobacterium tumefaciens strain GV3101 strain and then transiently expressed the CaABR1 gene in pepper plants using the same. It is a view showing the ion leakage results according to the death, active oxygen generation and cell death,

FIG. 7 shows the results of confirming the expression of disease-related genes of pepper plants through real-time PCR when CaABR1 gene was transiently overexpressed in pepper plants (CaBPR1: Capsicum annuum basic pathogenesis-related protein 1 , CaPR10: Capsicum annuum pathogenesis-related protein 10, CaDEF1: Capsicum annuum defensin 1),

8 is a result of confirming callus formation as a defense mechanism occurring in pepper plants when CaABR1 gene is transiently overexpressed in pepper plants through aniline blue staining,

9A and 9B show a recombinant vector TRV: CaABR1 prepared using a TRV vector and introduced into the Agrobacterium tumefaciens strain GV3101 strain, which was inoculated when Cayenne pepper plants were inoculated with CaABR1. This is a graph showing the plant photos showing the symptom and the pathogen growth in the plants after treatment with epsilic acid and inoculation of pathogenic strains of the red pepper bacterial spot pathogen.

Figures 10a to 10c is a trypan blue staining the plant photos and the plants showing the symptoms after treatment with epsis acid and inoculated non-pathogenic strains of red pepper bacterial spots bacteria produced CaABR1 gene silencing plants prepared as shown in Figures 9a and 9b (Trypan blue staining) photographs, and the result of the test of pathogen growth in the plant and the graph of the result of confirming the ion leakage,

11 is a recombinant vector of the pBIN35S: CaABR1 with Agrobacterium tumefaciens figure determined in transgenic Arabidopsis thaliana plants that overexpressed the CaABR1 gene using the (Agrobacterium tumefaciens strain GV3101) strains over-expressing if the CaABR1 gene by RT-PCR ego,

Figure 12 is the recombinant vector pBIN35S: CaABR1 and Agrobacterium tumefaciens (Agrobacterium tumefaciens strain GV3101) Pseudomonas when in transgenic Arabidopsis thaliana plants that overexpressed the CaABR1 genes using the strains ringge lost bar tomato DC3000 (Pseudomonas syringae pv. tomato DC3000) is a graph showing the results of assaying pathogen growth after inoculation,

FIG. 13 is a diagram showing the results of pathogenic resistance after inoculation of Hyaloperonospora parasica Noco2 ( Hyaloperonospora parasitica Noco2), which is a Bacillus bacterium, to a transgenic Arabidopsis plant that overexpresses the CaABR1 gene.

14 is a test for sporangiophore generated on leaves when inoculated with a bacterium hyaluroferon spora parasitia Noco2 ( Hyaloperonospora parasitica Noco2) in transgenic Arabidopsis plants overexpressing the CaABR1 gene. A graph showing one result,

15 is a conidia of spores generated from sporangiophore produced when disease resistance appears after inoculation of Hyalperonospora parasitica Noco2 ( Hyaloperonospora parasitica Noco2) as a bacterial bacterium to a transgenic Arabidopsis plant overexpressing the CaABR1 gene. (Conidiospore) is a graph showing the results of the test.

<110> Korea University Industrial and Academic Collaboration Foundation <120> Pepper disease resistance- and cell death-related, abscisic          acid-responsive gene CaABR1 and transgenic plants using the same <130> P11-080929-01 <160> 2 <170> KopatentIn 1.71 <210> 1 <211> 1172 <212> DNA 213 Capsicum annuum cv. Nockwang <400> 1 cttgaaaata gttttccgca cttaaatcag ttaatcatga caggcacaac agaagaaaat 60 caacccaaag ttcaagaatc agagcctcaa gcaccctcta ttcctacatc ttcttcttct 120 gatgagaaag agaaacaacc agaaatggat caacaaaaat ggggcacaca cataatgggt 180 ccaccagcag ttccaacaag tcatccagat aatcagaaag ctgctgcgtt atggagagct 240 gcagaccaaa aagaagagtt tcacccacag ccttacgttg tttatactcc agttgatcat 300 aggcctacta ataatccact tgaatctgtt gttaatatgt ttaattcttg gagtaatcga 360 gctgagacca tcgcccgcaa catctggcat aatctgagaa ctggaccatc agtgacagaa 420 gcagcgtggg gaaagcttaa tttgacagcc aaggccttaa cagaaggcgg attcgagccg 480 ctttacaagc agattttctc tacggaccct aatgagcagc tgaagaagac atttgcttgt 540 tatctttcaa caactactgg tcctgttgct ggaacactct atttgtcatc tactaaggtt 600 gctttttgca gtgatcgacc tttatccttc aaagctccat caggtcagga ggcttggagc 660 tactacaagg tagcaatacc attgacaaac attgggacta tgaacccaat agtgatgaga 720 gagaatccac cagagaggta cattcagata gttacaatcg atggtcatga cttctggttc 780 atggggtttg tcaattttga gaaagcaaca catcatctcc ttgatgcctt gtctaatttt 840 aaggcccaac ctcctcatgt tggggaagtg ccacaaccag ctagtaacta ggaaaaaatg 900 gctctgtcat aacttattgg gctgcacact gctttacttt ttctattttc tttgtttgtt 960 cctgcattag atatatgtat tctctgtatg tatttttatt gcaacattat ttgtattata 1020 cacactgctt tagtttttat tttctctgtt taggagattg acatatatac tgtctgtatt 1080 tttattgtaa catttttgta ttagactctc gtgtgtattt ttattgtaac attatttata 1140 ttgtaaaaaa aaaaaaaaaa aaaaaaaaaa aa 1172 <210> 2 <211> 284 <212> PRT 213 Capsicum annuum cv. Nockwang <400> 2 Met Thr Gly Thr Thr Glu Glu Asn Gln Pro Lys Val Gln Glu Ser Glu   1 5 10 15 Pro Gln Ala Pro Ser Ile Pro Thr Ser Ser Ser Ser Asp Glu Lys Glu              20 25 30 Lys Gln Pro Glu Met Asp Gln Gln Lys Trp Gly Thr His Ile Met Gly          35 40 45 Pro Pro Ala Val Pro Thr Ser His Pro Asp Asn Gln Lys Ala Ala Ala      50 55 60 Leu Trp Arg Ala Ala Asp Gln Lys Glu Glu Phe His Pro Gln Pro Tyr  65 70 75 80 Val Val Tyr Thr Pro Val Asp His Arg Pro Thr Asn Asn Pro Leu Glu                  85 90 95 Ser Val Val Asn Met Phe Asn Ser Trp Ser Asn Arg Ala Glu Thr Ile             100 105 110 Ala Arg Asn Ile Trp His Asn Leu Arg Thr Gly Pro Ser Val Thr Glu         115 120 125 Ala Ala Trp Gly Lys Leu Asn Leu Thr Ala Lys Ala Leu Thr Glu Gly     130 135 140 Gly Phe Glu Pro Leu Tyr Lys Gln Ile Phe Ser Thr Asp Pro Asn Glu 145 150 155 160 Gln Leu Lys Lys Thr Phe Ala Cys Tyr Leu Ser Thr Thr Thr Gly Pro                 165 170 175 Val Ala Gly Thr Leu Tyr Leu Ser Ser Thr Lys Val Ala Phe Cys Ser             180 185 190 Asp Arg Pro Leu Ser Phe Lys Ala Pro Ser Gly Gln Glu Ala Trp Ser         195 200 205 Tyr Tyr Lys Val Ala Ile Pro Leu Thr Asn Ile Gly Thr Met Asn Pro     210 215 220 Ile Val Met Arg Glu Asn Pro Pro Glu Arg Tyr Ile Gln Ile Val Thr 225 230 235 240 Ile Asp Gly His Asp Phe Trp Phe Met Gly Phe Val Asn Phe Glu Lys                 245 250 255 Ala Thr His His Leu Leu Asp Ala Leu Ser Asn Phe Lys Ala Gln Pro             260 265 270 Pro His Val Gly Glu Val Pro Gln Pro Ala Ser Asn         275 280  

Claims (10)

An isolated gene ( CaBR1 ) having the nucleotide sequence of SEQ ID NO: 1 from Capsicum annuum L. cv. Nockwang. An isolated protein having the amino acid sequence of SEQ ID NO: 2 (CaABR1). A recombinant vector comprising the CaABR1 gene of claim 1. A recombinant vector pBIN35S :: CaABR1 prepared by inserting the CaABR1 gene according to claim 1 into pBIN35S . A plant transformed by the gene of claim 1 or the recombinant vector of claim 3. The plant of claim 5, wherein the plant is pepper or Arabidopsis. A method for detecting resistance of a plant comprising the step of confirming the expression of the CaABR1 gene according to claim 1 by Northern blot analysis by separating RNA from a plant inoculated with non-pathogenic bacteria of red pepper bacterial spot pattern disease. Separation of RNA from plants treated with Abscisic acid (Abscisic acid) and the Northern blot analysis of the method of claim 1, comprising the step of confirming the expression of the CaABR1 gene described in claim 1. Recombinant vector TRV2 :: CaABR1 prepared by inserting the CaABR1 gene according to claim 1 into a Tobacco Rattle Virus 2 vector, which is a vector for virus-induced gene silencing (VIGS). Pepper, wherein the recombinant vector described in claim 9 is introduced to silence the CaABR1 gene according to claim 1.

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