KR20120125835A - Primer Set for Detecting Pectobacterium wasabiae and Method for Detecting by Using the Same - Google Patents

Abstract

PURPOSE: A primer set for detecting Pectobacterium wasabiae causing soft rot is provided to quickly and accurately detect Pectobacterium wasabiae and to prevent soft rot. CONSTITUTION: A primer set for detecting Pectobacterium wasabiae contains a primer with nucleic acid sequences of sequence numbers 1 and 2. A method for detecting Pectobacterium wasabiae comprises a step of amplifying DNA isolated from a plant sample and a step of identifying the PCR products. A composition for detecting Pectobacterium wasabiae contains the primer set. A PCR kit contains the primer set.

Description

Primer Set for Detecting Pectobacterium wasabiae and Method for Detecting by Using the Same}

The invention horseradish yeonbubyeong the causative organism of Peg tobak Te Solarium of wasabi (Soft rot) (Pectobacterium wasabiae ) and a primer set for detection , a method for detecting pectobacterium wasabi using the same, a composition for detecting pectobacterium wasabi containing the same and a PCR kit.

Soft rot is a Pectobacterium wasabiae ) is caused by a bacterium. Pectobacterium wasabi is a Gram-negative bacterium that is a causal anaerobic and causative agent of horseradish soft disease that produces pectate degrading enzymes.

Soft disease most frequently occurs during high temperatures in July and August, when the water temperature is 18 ° C or higher. As the roots collapse and collapse, they turn gray and odor, and yellowing of the leaves is a characteristic of soft disease.

There is no known method for controlling soft disease. In order to minimize the incidence, it is necessary to reduce the invasion of pathogens in the plant, and to thoroughly manage hygiene and prevent the temperature from rising above 15 ℃. In addition, seedlings should be soaked in 1000 times of farming mycin for 10 hours before planting, and harvested before passing twice in summer.

As such, there is a lack of measures for controlling harmful plant soft-pathogens, and thus, an accurate method for identifying and diagnosing pathogens is urgently needed.

Therefore, the inventors of the present invention, while repeatedly researching the effective detection method of wasabi soft disease causal bacteria, by using a method for identifying the species at the gene (DNA) level was to quickly and accurately diagnose wasabi soft disease.

Therefore, the present invention is a primer set that can effectively detect the bacterium bacterium wasabi bacterium at the gene level, a method for detecting the pectobacterium wasabi using the same, for detecting the pectobacterium wasabi containing the same It relates to a composition and a PCR kit.

In order to solve the above problems, the present invention is a Pectobacterium wasabi ( Pectobacterium) consisting of a primer having a nucleic acid sequence of SEQ ID NO: 1 and a primer having a nucleic acid sequence of SEQ ID NO: 2 wasabiae ) provides a primer set for detection.

In another aspect, the present invention includes the step of PCR amplifying DNA extracted from a plant specimen using a primer set consisting of a primer having a nucleic acid sequence of SEQ ID NO: 1 and a primer having a nucleic acid sequence of SEQ ID NO: 2 to confirm the presence of an amplification product. It provides a method for detecting pectobacterium wasabi.

In another aspect, the present invention provides a composition for detecting pectobacterium wasabi containing a primer set consisting of a primer having a nucleic acid sequence of SEQ ID NO: 1 and a primer having a nucleic acid sequence of SEQ ID NO: 2.

In another aspect, the present invention provides a PCR kit for detecting the pectobacterium wasabi containing a primer set consisting of a primer having a nucleic acid sequence of SEQ ID NO: 1 and a primer having a nucleic acid sequence of SEQ ID NO: 2.

By using the primer set according to the present invention, it is possible to quickly and accurately detect the bacterium bacterium wasabi, which is the causative agent of wasabi soft disease, and to effectively diagnose whether or not horseradish soft disease is infected, to prevent and control wasabi soft disease. Can contribute.

1 is a photograph showing the results of performing a PCR reaction using a primer set according to the present invention. (M: size marker, 1-34: strain of the number shown in Table 1, 35: control)
Figure 2 is a photograph showing the results of performing a real-time PCR reaction using a primer set according to the present invention. (1: genomic DNA of LMG 8444 in Pectobacterium wasabi, 2 to 5: horseradish leaves infected with Pectobacterium wasabi, 6 to 9: whole wasabi leaf control, 10: no template control)

The present invention is a Pectobacterium wasabi consisting of a primer having a nucleic acid sequence of SEQ ID NO: 1 and a primer having a nucleic acid sequence of SEQ ID NO: 2 ( Pectobacterium) wasabiae ) provides a primer set for detection.

Primers having the nucleic acid sequence of SEQ ID NO: 1 or SEQ ID NO: 2 are forward (PW7011F) and reverse (PW7011R) primers for amplifying 140 bp fragments of the YD repeat protein sequence of Pectobacterium wasabi, respectively.

The inventors of the present invention can search for nucleic acid sequence information capable of specifically detecting Pectobacterium wasabi using GenBank and blastn programs of the National Center for Biotechnology Information (NCBI) and detect the specific nucleic acid sequence. Primers were prepared. As a result of performing general PCR and real-time PCR amplification using the prepared primer set, it was confirmed that the primer set according to the present invention can specifically detect only Pectobacterium wasabi.

Pectobacterium wasabi is the causative agent of soft rot. By using the primer set according to the present invention, it is possible to specifically detect Pectobacterium wasabi, which is the causative agent of wasabi soft disease, and by using this, it is possible to accurately diagnose the infection of wasabi soft disease of the plant specimen to be diagnosed. have.

Therefore, the present invention includes a step of PCR amplifying DNA extracted from a plant specimen using a primer set consisting of a primer having a nucleic acid sequence of SEQ ID NO: 1 and a primer having a nucleic acid sequence of SEQ ID NO: 2 to confirm the presence of an amplification product. It provides a method for detecting pectobacterium wasabi.

As a result of PCR amplifying DNA extracted from a plant sample using the primer set according to the present invention, if a nucleic acid amplification product specific to pectobacterium wasabie is present, the plant sample can be diagnosed as was infected with horseradish soft disease.

In the present invention, "plant sample" refers to a target plant to be checked for infection with Pectobacterium wasabi, for example, cabbage, radish or horseradish, preferably horseradish, but is not limited thereto. It is not.

The method of extracting DNA from plant specimens can be carried out by conventional methods known in the art, which can be appropriately selected by those skilled in the art.

In one embodiment of the present invention, the PCR amplification may use a conventional PCR amplification reaction, preferably a conventional PCR amplification (conventional PCR) or real-time PCR amplification (real-time PCR) In case of using real-time PCR amplification reaction, quantitative analysis is possible in real time. Conventional PCR amplification reaction refers to a conventional PCR amplification reaction to perform a step of amplifying a specific DNA after amplifying a specific DNA, the step of confirming the amplification of the specific DNA is electrophoresis, etc. It can be carried out by checking the size of the DNA product amplified through. Specific PCR amplification methods can be appropriately selected and performed by those skilled in the art.

In another aspect, the present invention provides a composition for detecting pectobacterium wasabi containing a primer set consisting of a primer having a nucleic acid sequence of SEQ ID NO: 1 and a primer having a nucleic acid sequence of SEQ ID NO: 2.

The composition of the present invention includes a variety of reagents, such as PCR reaction mixtures, restriction enzymes, agarose, hybridization or electrophoresis, which are necessary for confirming experimental procedures and results necessary for detecting pectobacterium wasabi using the primer set. Necessary buffer solution may be further included.

More specifically, the present invention provides a PCR kit for detecting a pectobacterium wasabi containing a primer set consisting of a primer having a nucleic acid sequence of SEQ ID NO: 1 and a primer having a nucleic acid sequence of SEQ ID NO: 2.

The PCR kit for detection according to the present invention includes a reaction buffer solution, a polymerase, a dNTP, a stabilizer and all biological or chemical reagents, instructions for use, etc., in addition to the primer set to detect the pectobacterium wasabi in the sample. can do. Other configurations of such kits may be appropriately selected by those skilled in the art.

BEST MODE FOR CARRYING OUT THE INVENTION Hereinafter, the present invention will be described in detail with reference to the following examples. However, the following examples are intended to illustrate the contents of the present invention, but the scope of the present invention is not limited to the following examples. Embodiments of the present invention are provided to more fully describe the present invention to those skilled in the art.

< Example  1> Experimental strain

Other microorganisms, including Pectobacterium wasabiae , used in the present invention are the Belgian Co-ordinated Collections of Micro-organisms (BCCMTM) of Belgium and the Korean Agricultural Culture Collection (KACC) of the National Institute of Agricultural Science. And their sources are listed in Table 1.

Pectobacterium strains were cultured in Nutrient Agar medium (Meat extract 0.1%, Yeast extract 0.2%, Peptone 0.5%, Sodium chloride 0.5% Agar 1.5%: 1L).

number Strain name Deposit number source One Pectobacterium wasabiae LMG 8444 ^T Japan 2 Pectobacterium wasabiae KACC 10406 3 Pectobacterium wasabiae KACC 14078 Republic of Korea 4 Pectobacterium wasabiae KACC 10868 Japan 5 Pectobacterium wasabiae KACC 10869 Japan 6 Pectobacterium carotovorum subsp. carotovorum LMG 2404 ^T Denmark 7 Pectobacterium carotovorum subsp. carotovorum LMG 2399 UK 8 Pectobacterium carotovorum subsp. carotovorum LMG 2400 Japan 9 Pectobacterium carotovorum subsp. carotovorum LMG 2401 UK 10 Pectobacterium carotovorum subsp. carotovorum LMG 2405 Israel 11 Pectobacterium carotovorum subsp. carotovorum LMG 2406 Tanzania 12 Pectobacterium carotovorum subsp. carotovorum KACC 10343 Republic of Korea 13 Pectobacterium carotovorum subsp. carotovorum KACC 10347 14 Pectobacterium carotovorum subsp. carotovorum KACC 13968 15 Pectobacterium carotovorun subsp. odoriferum KACC 13954 16 Pectobacterium atrosepticum KACC 10477 ^T UK 17 Pectobacterium betavasculorum LMG 2466 ^T UK 18 Pectobacterium cacticida LMG 2720 USA 19 Erwinia rhapontici LMG 2688 ^T UK 20 Erwinia persicina KACC 10053 ^T Japan 21 Erwinia tracheiphila KACC 10084 ^T USA 22 Erwinia chrysanthemi KACC 10062 ^T USA 23 Erwinia herbicola LMG 2565 Canada 24 Erwinianigrifluence LMG 2694 ^T Zimbabwe 25 Enterobacter agglomerans LMG 1286 ^T USA 26 Pantoea ananatis LMG 2676 USA 27 Xanthomonas oryzae pv. oryzae KACC 10331 Republic of Korea 28 Pseudomonas syringae pv.syringae LMG 5082 UK 29 Pseudomonas fuscovaginae LMG 2158 ^T Japan 30 Pseudomonas graminis KACC 10803 ^T Germany 31 Rhizobium radiobacter KACC 10736 ^T 32 Ralstonia solanacerum KACC 10814 ^T USA 33 Serratia marcescens KACC 11961 ^T Czech Republic 34 Esherichia coli LMG 2092 ^T Denmark

^T , Type strain

< Example  2> primer  Set production

Example 2 -One: DNA  extraction

The total DNA of the strains cultured in Example 1 was extracted by the CTAB method, the total DNA of other microorganisms was extracted using a genomic DNA extraction kit (Genomic-tips) of Qiagen (Hilden, Germany).

Example 2 -2: singular Nucleic acid sequence  Information navigation

Pectobacterium wasabie, registered with GenBank (www.ncbi.nlm.nih.gov/) of the National Center for Biotechnology Information (NCBI) for the preparation of primer sets for specifically detecting pectobacterium wasabies Genomic information of the blastn program was analyzed by the sequence information (SEQ ID NO: 3) of the YD repeat protein was confirmed to be specific.

Example  2-3: primer  Set production

PW7011F primer and PW7011R primer set were prepared to amplify 140 bp fragment (SEQ ID NO: 4) from YD repeat protein (SEQ ID NO: 3) of Pectobacterium wasabi. The primer sequences are shown in Table 2 below.

SEQ ID NO: designation order One PW7011F 5'-ctatgacgctcgcgggttgctgtt-3 ' 2 PW7011R 5'-cggcggcgtcgtagtggaaagtc-3 '

The primer was prepared after confirming specificity of the sequence information of GenBank YD repeat protein through a blastn program.

< Example  3> primer  Confirm the specificity of the set

Example 3 -1: normal PCR  Amplification reaction

PCR amplification reaction was performed on the strains described in Table 1 using the primer set consisting of SEQ ID NO: 1 (PW7011F) and SEQ ID NO: 2 (PW7011R).

PCR amplification reaction was carried out using PTC-200TM thermocycler (MJ research, Watertown, mass), each reaction was Tris-HCl 10mM, KCl 50mM, MgCl2 1.5mM, gelatin 0.01%, dNTP each 0.2mM, primers 10pM, A PCR mixture containing 2 units of Tac polymerase (Taq polymerase, Promega, Madison, Wis.) Was performed in a total amount of 50 μl. The amount of genomic DNA of the strains added to the PCR mixture was about 50ng.

The reaction of Pectobacterium wasabi was denatured at 94 ° C. for 5 minutes, repeated 40 times at 94 ° C. for 60 seconds, at 67 ° C. for 30 seconds, and at 72 ° C. for 60 seconds and then at 72 ° C. for 10 minutes. After the amplification reaction, 10 μl of PCR amplification product was electrophoresed on agarose gel at 1.5% concentration, and then stained on ethidium bromide, which was confirmed by a UV transilluminator, and the results are shown in FIG. 1.

In Figure 1, M is a DNA sizing marker (1kb ladder, Gibco BRL ^TM ), the numbers 1 to 34 means the strain of the number shown in Table 1, 35 is a control using distilled water. As can be seen from Figure 1, the band showing the PCR amplification product was confirmed only in the 1 to 5 Pectobacterium wasabi strain bar, primer set according to the present invention specifically pectobacterium wasabi strain It can be seen that it can be detected.

Example 3 -2: Real - time PCR  Amplification reaction

Real-time PCR amplification reactions were performed using the CFX96 real-time PCR system (Bio-Rad laboratories, Inc., USA), each reaction using a KAPA SYBR®FAST Universal 2X qPCR Master Mix (KAPA Biosystem, Inc., USA). A total of 20 μl was performed with a PCR mixture containing 10 μl and 5 pM each of primers. The amount of genomic DNA of pectobacterium wasabi added to the real-time PCR mixture was about 5ng, and extracted directly from 1cm x 1cm horseradish tissues infected with wasabi, and added 1µl each to Real-time Bio-PCR. Was done.

The reaction of Pectobacterium wasabi was denatured at 95 ° C. for 2 minutes 30 seconds, repeated 45 times at 95 ° C. for 10 seconds, and at 67 ° C. for 20 seconds, and then denatured at 95 ° C. for 10 seconds and 0.5 ° C. at 65 ° C. to 95 ° C. Checking the melt curve and the melt peak while raising. The results are shown in Fig.

2 shows genomic DNA of LMG 8444 in Pectobacterium wasabi, 2 to 5 horseradish leaves infected with Pectobacterium wasabi, 6 to 9 whole horseradish leaf controls, and 10 to no template controls. In FIG. 2A, the curve portion at the front is genomic DNA of LMG 8444 at Pectobacterium wasabi and the curve at the back is at Wasabi tissue (2-5) infected with wasabi soft bacilli (Pectobacterium wasabi). Specific PCR fragments present in common are shown. Nucleic acid fragments were specifically amplified only in horseradish tissues infected with Pectobacterium wasabi LMG 8444 genomic DNA and horseradish soft germ (Pectobacterium wasabi).

Therefore, a primer set consisting of a primer having a nucleic acid sequence of SEQ ID NO: 1 (PW7011F) and a primer having a nucleic acid sequence of SEQ ID NO: 2 (PW7011R) can amplify only a specific nucleic acid fragment of horseradish soft germ bacterium. It can be seen that it can be used as a PCR primer for diagnosis of infection, and confirmed that it can be specifically detected in 1cm x 1cm tissue infected with horseradish soft disease.

Having described the specific parts of the present invention in detail, it will be apparent to those skilled in the art that such specific descriptions are merely preferred embodiments, and thus the scope of the present invention is not limited thereto. will be. Accordingly, the actual scope of the present invention will be defined by the appended claims and their equivalents.

<110> Rural Development Administration <120> Primer Set for Detecting Pectobacterium wasabiae and Method for          Detecting by Using the Same <130> P110368 <160> 4 <170> Kopatentin 1.71 <210> 1 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> primer for detecting Pectobacterium wasabiae <400> 1 ctatgacgct cgcgggttgc tgtt 24 <210> 2 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> primer for detecting Pectobacterium wasabiae <400> 2 cggcggcgtc gtagtggaaa gtc 23 <210> 3 <211> 4140 <212> DNA <213> Pectobacterium wasabiae <400> 3 atgtttgaag ctgcgcgcgt tggcgatgga ataggacact ccagtgctct gactggcatg 60 atcctgggca ccatcgctgg cggattgatt gctgccgccg gcggtattgc ggccggtgct 120 ctgcttgtgg cggggctggg ctcctgtgtg gtgggtggcg tgttactgat tggcctcagt 180 gctgcggtgg ggtgggcgac gggcgagctg gcagaaaagg ttcgtgacgg cattgccgac 240 tcctttgcca gcagcatgtc aaaggccggc acgattgacg agggctcacc cacggtcttt 300 atcaacggca tgaaggccgc catggctatc ctgagtatag ccagttgcaa taaagacggc 360 ccttccatgc aggtggcgga gggctccgca acggtctaca tcaacaacca gcccgcctcc 420 cgtctggggg acaaggtgaa ctgcggtgcc actatcacag aaggctcatc caatgtgttc 480 attggggggg cgacgaaacc catgctgcca atccggtcag aagtgccggc ctggctctat 540 aaagcctctg atttaacgct gcttttcgcc gggctggttg gcggggtggg gggcatggcg 600 ggcaaagccg gaaaactggg taagctgctg cgggaacttc ccggtctcaa caagctccag 660 cgtcttgcct gccgttttgg cgtactcatg accagtatcg ccgccggggg gattattgcc 720 cgcccaatcg acatcgtgag cgggcaaaaa tttctggcgg atgaggagga gctggatttc 780 gtcctacctt cccgcttgcc cgtctattgg caacgctgct ggcgtagcgg caatcccggt 840 gatagcgtat tgggaaaggg atggagcctg ttctgggaaa cgacactgac ctgctatcag 900 gacggtctgg tctggcgcgc cccgtcaggc gatctgattt cctttccttt tgttcccgtc 960 ggccagcgaa gctactgtcc ggcagaaaag cgctggctgg agcaccatca ggatgaaagc 1020 tggtcggttt atgggccgga cggcgaagta tggtcttaca ccgctttgtc tccacagggg 1080 aaagcggttc tggcacgcat tgccgaaccg tgtggtcatg acattctctt tttctggaac 1140 gatgatgaca cgcttcaggc gctaacggat agcgccggac ggcacctcac ctgccggtat 1200 cgtgacgggc gactggacag tgtctggctg gatgaatcga cctgtctggt gcactatacc 1260 tacaatgcgc aaaggcagtt gattaccgtc accgggcggg gcggtagcgt gcgccgtcgt 1320 tttacttggc acgatgacgg gctgatggcc agccatgaag aggcgagcgg cctactcagc 1380 gaatatcagt ggcaggagat tgccgattta ccgcgtgtgg tggcctaccg caacagcgcg 1440 ggtgagcagc ttacgctgga gtatgatttc gccggacaac ggcgaaccgc ccgccgagag 1500 gatggtctca tcgcgcagtg gttacttgac gacgaaggtc acgtcactca ctttaccgat 1560 tacgacggcc gtgagaccac tctgagttat gccgatggcg aactgtgtga cgtcatcatg 1620 cccggtgggg cgagccgcaa aacgacctgg gacgattatg gtcgtttact cagcgagacc 1680 gatccgctgg gtcgggaaac ttgctaccag tggtatcggc tgacgggtca cattacgctg 1740 gtcacgtatc ccgacggcag ccgcgagcaa atgcagtatg acgacctcaa tcgtctggtt 1800 gaggaaattg acgcacgggg taactcaact cgctaccagt accccacggc acaggaaagt 1860 ttgccggaca gcatcaccga cgcactgggc ggcaccgtca cgctggtctg gaaccgacag 1920 gggctgctta cggcacgtac cgactgttcc gggcagtgca gcacctttga gtatgaccat 1980 gacgggcagt tgctggcatc tgttgatgcc gaaggtcaca ccacccgccg ggaatgggat 2040 gcctgcgggc acctgacaag cgttatctac ccggatggtc gccgggaaac actgcgctgg 2100 aatgcccggg gtcagttgca ggcctggcgt gacgcacaga acagcgaagt ccgctggcac 2160 tataacgtcc tcggccagcc cgtcagtgta acggaccgtc ttcgccggac aacgcgctgg 2220 cactatgacg ctcgcgggtt gctgttacgg ctggagaacg gcaacggcgc agcgtaccag 2280 tttggctatg atgccgtcgg ccgactggtt aacgagcagc gcgtggatgg cgtcgagctg 2340 actttccact acgacgccgc cggtcacctg tgccagcggt cgcagcgcgg tcatgccgcg 2400 aatgacgaag ctatctcaca catttatcag cacgatgcgg ccgggcaact ggtgcgtcgt 2460 gaacatgccc acgccgtgtt ccactaccag cacaacaaca gagggcaact gctttccctg 2520 aagcgtgaac caaccgggga aggcctcgcg ctgggtctca ccgccgatga ggtcaggctg 2580 acctatgacg cagcgggcga cctgtcaggc gaacacggca gtgaaggcga catcggctat 2640 acccgggatg cactgggcaa cgtgagtgcc ctgtcgctac cggatggtga cacgctgcgc 2700 tggttgcgct acggctccgg ccacgtcagc gcggtaaaat tcaaccatca ggtggtcagc 2760 gagtttaccc gtgaccggtt gcaccgggaa atcagccgta ctcagggccg ccgcacgcag 2820 caacgggcgt atgacagcct cggtcgcctg acctcgcagc gcagcggtct ctgggatgtc 2880 gcggagccgg agcagcaact tctctcgcgg gcgttacgct acacggcttc cggcgaactc 2940 gcttcagtca gggacagcct gcggggtgac gtgcagtatg attacgatgc agaaggtcgc 3000 ctgctgaaac ggattgacgt gcactggcag gtgcatcatc gggcatatgg ctatgatgcg 3060 gctgacaatc tgcaggatag cgcttattca ccgtctgccc gcccactctc cgacaaccgc 3120 ctgctgaact ggcgtcatct gtggaaccag tatgacggcc agggcaacct gatacggcgg 3180 cgtgaaggca caacggagca gttttaccag tatgatgcgg acaaccgtct ggttgaagcc 3240 cggggccggg ggccacaggg cgagtttgtg gcccgctatg gctatgatgc gctgggcaga 3300 cgcacccgta aaacggtcac ctggggtgac agcgggaaac aggaagagac gcgtttcctg 3360 tgggaaggtt ttcggctgtt gcaggccaga caggctgacc ggacagaaac ctacctttat 3420 gacccgacta tctggtggtc gccgttagca cgtatcacac agcaaccggg cgcgcctgac 3480 ggtgatatcc gctggttcaa taccgagctg aacggcgcgc cgctggagat gacggatgcg 3540 gaaggtgcgg tccgctggag tggagattac ggcagcttcg gtgccatcaa tggacagacg 3600 caggacagtg aaggactgcg ccacggcaag ccggtagaat cccagtcgct acgctatgcc 3660 ggacaatacg cggatgagga aacggggtta cactacaacc tgttccgcta ttacgacccc 3720 accgtaggcc gcttcacgac acaggacccg ataggcctgg cgggagggct gaatctttat 3780 gcttatgcgc cgaatccgct ggggtgggtg gatccgctgg ggttggccaa aatattaacc 3840 gatggtgttg tctatcgtat gggaagtgga accgactcaa atttgacccc tcgaccagga 3900 aaagatacaa cgacaggact atctacaact atcgagaaac cttctaatgg taaatatcaa 3960 actcttgatg tgaaaacgct aaatgatggc ggattagatg tagttaaaga tggtaaaaat 4020 catgcttctg tcaggccaag taatgatcca gatatgtcta gattgagaga atgggctgat 4080 accagaggaa ccgagaaatc gtcttcttat acaaagactg taaagaaatc atgtggataa 4140                                                                         4140 <210> 4 <211> 140 <212> DNA <213> Pectobacterium wasabiae <400> 4 ctatgacgct cgcgggttgc tgttacggct ggagaacggc aacggcgcag cgtaccagtt 60 tggctatgat gccgtcggcc gactggttaa cgagcagcgc gtggatggcg tcgagctgac 120 tttccactac gacgccgccg 140

Claims (5)

A primer set for detecting Pectobacterium wasabiae consisting of a primer having a nucleic acid sequence of SEQ ID NO: 1 and a primer having a nucleic acid sequence of SEQ ID NO: 2.
PCR amplification of DNA extracted from plant specimens using a primer set consisting of a primer having a nucleic acid sequence of SEQ ID NO: 1 and a primer having a nucleic acid sequence of SEQ ID NO: 2 to confirm whether amplification product is present. Detection method of Leeum wasabi.
The method of claim 2,
The plant sample is a method for detecting pectobacterium wasabi that was wasabi.
A composition for detecting pectobacterium wasabi comprising a primer set consisting of a primer having a nucleic acid sequence of SEQ ID NO: 1 and a primer having a nucleic acid sequence of SEQ ID NO: 2.
PCR kit for detecting a pectobacterium wasabi containing a primer set consisting of a primer having a nucleic acid sequence of SEQ ID NO: 1 and a primer having a nucleic acid sequence of SEQ ID NO: 2.

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