KR101444219B1 - Method of identifying Xanthomonas melonis - Google Patents

Abstract

The present invention relates to a method for identifying Xanthomonas melonis, and more particularly, to a method for identifying a base of 23S rRNA isolated from a sample to identify Xanthomonas melonis. When the identification method of Xanthomonas melonis of the present invention and the probe for identification of Xanthomonas melonis are used, it is possible to effectively identify Xanthomonas melonis from Xanthomonas have.

Description

{Method of identifying Xanthomonas melonis}

The present invention relates to a method for identifying Xanthomonas melonis, and more particularly, to a method for identifying a base of 23S rRNA isolated from a sample to identify Xanthomonas melonis.

The genus Santomonas (Xanthomonas) is a bacteria involved in the denitrification of soil nitrogen. These include pathogenic bacteria such as peach tree bacterial pericarditis, non-black rot, citrus peptic ulcer disease, rice blight white leaf blight,

Xanthomonas melonis is a proteobacteria species. These plant pathogens cause cucumber and melon disease.

Dot-blot, PCR (PCR), etc. are used for gene identification of plant pathogenic bacteria. In particular, PCR is a widely used method because it is economical in terms of time, effort, and manpower than the dot-blot method. PCR (polymerase chain reaction) is a method for identifying a DNA fragment (primer) consisting of 10-20 bp in the genomic DNA of a bacterium and then using heat-resistant DNA polymerase And the synthesis reaction is repeatedly carried out, the region to which the primer is bound is rapidly amplified. The PCR amplification product is electrophoresed on agarose gel or acrylamide gel, and ethidium bromide

the DNA polymorphism of bacterial species such as the presence or absence of DNA bands after DNA staining with ethidium bromide and silver stain is identified and identified. At present, the identification method of bacteria by PCR is mainly used for identification of bacterial diseases in humans and animals, but in recent years, PCR identification methods have been developed to identify plant pathogens

It is used for identification of illness and for quarantine of import and export agricultural products. In addition, the PCR identification method is shorter in time than other methods, is low in cost for identification, and is also economical because many samples can be assayed at the same time. Recently, PCR identification method that amplifies nucleic acid which is a genetic material among the identification methods of Santomonas (Xanthomonas) is most preferred.

However, existing PCR identification methods are difficult to classify due to genetic similarity, and a solution to overcome this problem is needed.

Among the existing microorganisms of the genus Santomonas, Xanthomonas oryzae pv. Oryzae, Xanthomonas campestris pv. Vesicatoria, Xanthomonas campestris pv. Campestris, Xanthomonas axonopodis pv. citr and Xanthomonas axonopodis pv. glycines. However, it has been reported that the PCR primers used for the identification of Xanthomonas melonis from Xanthomonas The method has not yet been studied. There is a need to develop a method capable of identifying Santomonas melonis through the combination of species or pathogen-specific bases and bases between plant pathogenic bacteria.

Accordingly, the inventors of the present invention discovered the difference in the 23S rRNA sequence of the bacteria of the genus Santomonas during the study for more accurate and rapid identification of Xanthomonas melonis from Xanthomonas We developed a method for identification.

Accordingly, an object of the present invention is to provide a method for detecting (i) isolating 23S rRNA from a sample;

(b) confirming the nucleotide sequence of the 23S rRNA isolated in the step (a); And

(c) the nucleotide sequence identified in step (b) is the 78th base G of SEQ ID NO: 1, the 86th base C of SEQ ID NO: 1, the 682th base C of SEQ ID NO: 1, the 740th base A of SEQ ID NO: Base 1177 of SEQ ID NO: 1, nucleotide T at position 1357 of SEQ ID NO: 1, nucleotide T of nucleotide 1735 of SEQ ID NO: 1, nucleotide 1966 of base 1966 of SEQ ID NO: 1, nucleotide 1967 of nucleotide 1967 of SEQ ID NO: 1, nucleotide 1969 The nucleotide 1973 of SEQ ID NO: 1, the nucleotide 1977 of SEQ ID NO: 1, the nucleotide 1978 of SEQ ID NO: 1, the nucleotide 1979 of SEQ ID NO: 1, the nucleotide 2048 of SEQ ID NO: 1, The 2049th base G of SEQ ID NO: 1, the 2053th base G of SEQ ID NO: 1, the 2056th base A of SEQ ID NO: 1, the 2058th base A of SEQ ID NO: 1, the 2060th base G of SEQ ID NO: Base A at position 2061 of SEQ ID NO: 1, nucleotide 2626 of base 262 of SEQ ID NO: 1, base 2649 at nucleotide 2649 of SEQ ID NO: 1, 2715th base C of SEQ ID NO: 1, and Xanthomonas melonis in the case of 2737th base G of SEQ ID NO: 1.

Another object of the present invention is a polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 78th base G of SEQ ID NO: 1, 20-100 consecutive DNAs comprising the 86th base C of SEQ ID NO: A polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 682nd nucleotide C of SEQ ID NO: 1, 20-100 consecutive DNAs comprising the 740th nucleotide A of SEQ ID NO: 1 A polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 1177th nucleotide C of SEQ ID NO: 1, 20-100 consecutive DNAs comprising the 1357th nucleotide T of SEQ ID NO: 1 A polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 1735th base T of SEQ ID NO: 1, a polynucleotide consisting of 1966th base A of SEQ ID NO: 1, A polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 1967th base T of SEQ ID NO: 1, a polynucleotide consisting of 20-100 consecutive DNA sequences comprising 1967th nucleotide T of SEQ ID NO: 1, a 1969th nucleotide C of SEQ ID NO: 1 A polynucleotide consisting of 20-100 consecutive DNA sequences comprising SEQ ID NO: 1, a polynucleotide consisting of 20-100 consecutive DNA sequences comprising 1973th nucleotide C of SEQ ID NO: 1, a polynucleotide consisting of 1977th nucleotide C of SEQ ID NO: 1 A polynucleotide consisting of 20-100 consecutive DNA sequences comprising SEQ ID NO: 1, a polynucleotide consisting of 20-100 consecutive DNA sequences comprising 1978th nucleotide T of SEQ ID NO: 1, a polynucleotide consisting of 1979th nucleotide C of SEQ ID NO: 1 A polynucleotide consisting of 20-100 consecutive DNA sequences comprising SEQ ID NO: 1, 20-100 consecutive DNA sequences comprising 2048th nucleotide G of SEQ ID NO: A polynucleotide consisting of 20-100 consecutive DNA sequences comprising 2049th nucleotide A of SEQ ID NO: 1, 20-100 consecutive DNA sequences comprising 2050th nucleotide G of SEQ ID NO: 1, A polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 2053 base G of SEQ ID NO: 1, a polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 2056th base A of SEQ ID NO: 1, A polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 2058th base A of SEQ ID NO: 1, a polynucleotide consisting of 20-100 consecutive DNA sequences comprising 2060th base A of SEQ ID NO: 1, Nucleotide, a polynucleotide consisting of 20-100 consecutive DNA sequences including 2061th base T of SEQ ID NO: 1, 2626th base T of SEQ ID NO: 1 Comprises a polynucleotide consisting of 20-100 consecutive DNA sequences, a polynucleotide consisting of 20-100 consecutive DNA sequences comprising 2649th base T of SEQ ID NO: 1, 2715th nucleotide C of SEQ ID NO: 1 A polynucleotide consisting of 20-100 consecutive DNA sequences consisting of 20-100 consecutive DNA sequences comprising 2737th base G of SEQ ID NO: 1, and a complementary polynucleotide thereof And a probe for identifying Xanthomonas melonis comprising at least one selected polynucleotide.

In order to achieve the above object, the present invention provides a method for identification of Xanthomonas melonis.

In order to accomplish another object of the present invention, there is provided a probe for identification of Xanthomonas melonis.

Hereinafter, the present invention will be described in detail.

The present invention relates to a method for identification of Xanthomonas melonis.

That is, the present invention secures a base combination of the 23S rRNA gene of Xanthomonas to identify Xanthomonas melonis.

More specifically, the method for identifying Xanthomonas melonis of the present invention comprises the steps of: (a) separating 23S rRNA from a sample; (b) confirming the nucleotide sequence of the 23S rRNA isolated in the step (a); And (c) the nucleotide sequence identified in step (b) is the 78th base G of SEQ ID NO: 1, the 86th base C of SEQ ID NO: 1, the 682th base C of SEQ ID NO: 1, the 740th base A , Nucleotide 1177 of SEQ ID NO: 1, nucleotide T at position 1357 of SEQ ID NO: 1, nucleotide T at nucleotide 1735 of SEQ ID NO: 1, nucleotide A at position 1966 of SEQ ID NO: 1, nucleotide 1967 of nucleotide 1967 of SEQ ID NO: 1, 1969th nucleotide C of SEQ ID NO: 1, 1977th nucleotide C of SEQ ID NO: 1, 1978th nucleotide T of SEQ ID NO: 1, 1979th nucleotide C of SEQ ID NO: 1, 2048th nucleotide G of SEQ ID NO: 1, The 2049th base G of SEQ ID NO: 1, the 2050th base G of SEQ ID NO: 1, the 2053th base G of SEQ ID NO: 1, the 2056th base A of SEQ ID NO: 1, the 2058th base A of SEQ ID NO: 1, A nucleotide T at position 2061 of SEQ ID NO: 1, a nucleotide T at position 2626 of SEQ ID NO: 1, a nucleotide T at position 2649 of SEQ ID NO: 1, a nucleotide sequence of SEQ ID NO: When the 2715th base C, 2737 beonjjae bases of SEQ ID NO: 1 G is characterized in comprising the step of determining a fitness Santo Pseudomonas melo (Xanthomonas melonis).

The present invention also provides a probe for identifying Xanthomonas melonis comprising a gene base combination.

That is, the present invention secures a base combination of the 23S rRNA gene of Xanthomonas to identify Xanthomonas melonis.

More specifically, the probe for identifying Santomonas melonis of the present invention comprises a polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 78th base G of SEQ ID NO: 1, a polynucleotide consisting of the 86th base of SEQ ID NO: 1 A polynucleotide consisting of 20-100 consecutive DNA sequences containing C, a polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 682nd nucleotide C of SEQ ID NO: 1, a polynucleotide consisting of 20 nucleotides A, a polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 1177th base C of SEQ ID NO: 1, a polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 1177th nucleotide C of SEQ ID NO: 1, a polynucleotide consisting of 20-100 consecutive DNA sequences comprising SEQ ID NO: T, a polynucleotide consisting of 20-100 consecutive DNA sequences containing T, a 20-100 consecutive DNA sequences containing 1735th base T of SEQ ID NO: 1 A polynucleotide consisting of 20-100 consecutive DNA sequences comprising 1966th base A of SEQ ID NO: 1, 20-100 consecutive DNA sequences comprising 1967th nucleotide T of SEQ ID NO: 1 A polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 1969th base C of SEQ ID NO: 1, and 20-100 consecutive DNA sequences comprising 1973th nucleotide C of SEQ ID NO: 1 A polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 1977th base C of SEQ ID NO: 1, 20-100 consecutive DNA sequences comprising 1978th nucleotide T of SEQ ID NO: 1 A polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 1979th base C of SEQ ID NO: 1, a polynucleotide consisting of 2048th salt of SEQ ID NO: 1 G, a polynucleotide consisting of 20-100 consecutive DNA sequences comprising 2049th base A of SEQ ID NO: 1, a polynucleotide consisting of 20-100 consecutive DNA sequences comprising 2049th base A of SEQ ID NO: 1, a polynucleotide consisting of 2050th base A polynucleotide consisting of 20-100 consecutive DNA sequences containing G, a polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 2053th base G of SEQ ID NO: 1, a polynucleotide consisting of 20-100 consecutive DNA sequences comprising SEQ ID NO: A, a polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 2058th base A of SEQ ID NO: 1, a polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 2058th base A of SEQ ID NO: 1, a polynucleotide consisting of 2060th base A polynucleotide consisting of 20-100 consecutive DNA sequences comprising SEQ ID NO: 1, 20-100 consecutive DNA sequences comprising 2061th nucleotides T of SEQ ID NO: 1 Comprises a polynucleotide, a polynucleotide consisting of 20-100 consecutive DNA sequences comprising 2626 base T of SEQ ID NO: 1, 20-100 consecutive DNA sequences comprising 2649th base T of SEQ ID NO: 1 A polynucleotide consisting of 20-100 consecutive DNA sequences comprising 2715th base C of SEQ ID NO: 1, 20-100 consecutive DNA sequences comprising 2737th nucleotide G of SEQ ID NO: 1 And a complementary polynucleotide of the polynucleotide and the polynucleotide of the present invention.

In the present invention, the nucleic acid may be separated in step (a) according to a method commonly used in the art, and may be carried out using a commercially available extraction kit.

Also, the base sequence in step (b) may be performed according to automatic or manual base sequence analysis methods known in the art. Preferably, PCR is performed on a test specimen or a sample to perform PCR corresponding to 23S rRNA And the sequence of the product can be analyzed and analyzed according to a known nucleotide sequence analysis method.

In the step (c), the nucleotide sequence corresponding to the judgment point in the microorganism of the present invention can be identified based on SEQ ID NO: 1 to determine whether the microorganism corresponds to the present invention.

Meanwhile, the present invention provides a microarray for detecting microorganisms of the present invention, wherein the probe of the present invention is integrated on a substrate. The microarray consists of a conventional microarray except that it contains a polynucleotide of the present invention, and a method for producing a microarray by immobilizing a polynucleotide used as a marker on an organ is well known in the art.

Further, the present invention is a marker for detecting a microorganism according to the present invention, wherein a nucleotide having a nucleotide sequence shown in SEQ ID NO: 1 is substituted for each of the bases corresponding to the judgment point of the microorganism of the present invention. Lt; / RTI > The above judgment points are as shown in Table 3.

The nucleotide, probe or marker of the present invention may be DNA or RNA, and it is obvious to those skilled in the art that the thymine described in the sequence is replaced by uracil in the case of RNA.

Therefore, when the identification method of Xanthomonas melonis of the present invention and the probe for identification of Xanthomonas melonis are used, it is possible to effectively identify Xanthomonas melonis from Xanthomonas genus can do.

Fig. 1 shows the result of comparative analysis of nucleotide sequences of Xanthomonas campestris pv. Campestris and Xanthomonas melonis.

Hereinafter, the present invention will be described in detail with reference to examples.

However, the following examples are illustrative of the present invention, and the present invention is not limited to the following examples.

< Example  1>

X. melonis  And Hospitals  23S rRNA  analysis

Through the following experimental procedure, a probe for identification of bacteria belonging to the genus Xanthomonas was designed. The total RNA isolation, base sequence amplification and analysis methods used in the present invention are as follows.

<1-1> 23S rRNA amplification of bacteria

After the DNA of X. theicola was isolated, PCR was performed using the primer pairs shown in Table 1 below, which can isolate 23s rRNA of bacteria of the genus Xanthomonas.

Name of the primer Primer sequence Xan23SF5 (omnidirectional) 5'-TGGTCAAGCCGCACGGATCATTAGTAT-3 ' Xan23SR6 (reverse direction) 5'-ACGTGGATAGCCTGCGAAAAGTGTC-3 ' Xan23SF7 (Omnidirectional) 5'-GAGACCGCCCCAGTCAAACTAC-3 ' Xan23SR7 (reverse direction) 5'-ACCTTTTGTATAATGGGTCAACG-3 ' Xan23SF9 (Omnidirectional) 5'-GTCAAGCCGCACGGATCATTAGTAT-3 ' Xan23SR9 (reverse direction) 5'-GTCGGGGAGCTGGCAACAAG-3 '

The PCR reaction consisted of 20 ng of isolated total DNA, 10 pmoles of downstream primer, 10 pmole of upstream primer, 0.2 U of Taq DNA polymerase, 10 times Polymerase chain reaction buffer (100 mM Tris-HCl, 500 mM KCl, 15 mM MgCl ₂ , pH 8.3) was added and distilled water was added to make 50 μl of reaction solution.

The reaction solution was heated at 95 ° C. for 5 minutes and then amplified 30 times at 95 ° C. for 30 seconds, 65 ° C. for 60 seconds, and 72 ° C. for 180 seconds. Finally, the reaction solution was treated at 72 ° C. for 10 minutes with a PCR machine (DNA Engine PTC-200) Respectively. After amplification, the PCR product was electrophoresed using 1 × TBE buffer and 1.5% agarose gel, stained with ethidium bromide, and then confirmed by ultraviolet lamp.

<1-2> Amplified 23s rRNA  Sequencing

The nucleotide sequences of the PCR products obtained in Example <1-1> were analyzed using BioEdit Sequence Alignment Editor.

After analysis, X. campestris pv. We compared the 23s rRNA sequences of campestris. X. campestris pv. For the campestris 23s rRNA sequence, the information (NC_003902.1) published in Genbank was used. The 23s rRNA sequences of the bacteria used in this experiment are summarized in the sequence numbers as shown in Table 2 below.

SEQ ID NO: Germ One X. campestris pv. campestris 2 X. melonis

< Example  2>

Interspecific  23s rRNA  Sequence comparative analysis

The nucleotide sequence differences between SEQ ID NOS: 1 and 2 in Table 2 were compared. The nucleotide sequences were compared using the BioEdit Sequence Alignment Editor.

The results of the analysis are shown in Fig. 1, and X. campestris pv. Table 3 summarizes the positions of nucleotide sequence differences with X. melonis based on the nucleotide sequence of Campestris (SEQ ID NO: 1).

Position in sequence number  base No. 78 of SEQ ID NO: 1 G The 86 th C No. 682 of SEQ ID NO: 1 C 740 &lt; th &gt; A No. 1177 of SEQ ID NO: 1 C No. 1357 of SEQ ID NO: T No. 1735 of SEQ ID NO: T The 1966th A The 1967th T The 1969th C The 1973 th C 1977 th in SEQ ID NO: 1 C 1978 in SEQ ID NO: 1 T 1979 th in SEQ ID NO: 1 C The 2048th G The 2049th A The 2050th G No. 2053 of SEQ ID NO: G No. 2056 of SEQ ID NO: A No. 2052 of SEQ ID NO: A The 2060th A The 2061th T 2626nd of SEQ ID NO: C The 2649th T 2715 th in SEQ ID NO: 1 C 2737 th in SEQ ID NO: G

Therefore, the bacterium can be identified using the difference of the bases shown in [Table 3] above.

As described above, when the identification method of Xanthomonas melonis of the present invention and the probe for identification of Xanthomonas melonis are used, Xanthomonas melonis is obtained from Xanthomonas It can be identified effectively, and thus the possibility of industrial use is high.

<110> REPUBLIC OF KOREA (MANAGEMENT: RURAL DEVELOPMENT ADMINISTRATION) <120> Method of identifying Xanthomonas melonis <130> NP12-0058 <160> 8 <170> Kopatentin 2.0 <210> 1 <211> 2883 <212> DNA <213> X. campestris pv. campestris NC_003902 <400> 1 atggtcaagc cgcacggatc attagtatca gttagctcaa tacattgctg tacttacaca 60 cctgacctat caaccacata gtctatatgg ttcctttagg gggcttgtgc cccgggaagt 120 ctcatcttga ggcgcgcttc ccgcttagat gctttcagcg gttatcgctt ccgaacatag 180 ctacccggca atgccactgg cgtgacaacc ggaacaccag aggttcgtcc actccggtcc 240 tctcgtacta ggagcagccc ctctcaaact tccaacgccc atggcagata gggaccgaac 300 tgtctcacga cgttctgaac ccagctcgcg taccacttta aatggcgaac agccataccc 360 ttgggaccga ctacagcccc aggatgtgat gagccgacat cgaggtgcca aacaccgccg 420 tcgatatgaa ctcttgggcg gtatcagcct gttatccccg gagtaccttt tatccgttga 480 gcgatggccc ttccatacag aaccaccgga tcactaagac ctactttcgt acctgcttga 540 tccgtcgatc ttgcagtcaa gcacgcttat gcctttgcac acagtgcgcg atgtccgacc 600 gcgctgagcg taccttcgtg ctcctccgtt actctttagg aggagaccgc cccagtcaaa 660 ctacccacca tacacggtcc ctgatccgga taacggatct aggttagaac gtcaagcacg 720 acagggtggt atttcaaggt tggctccact gcagctagcg ccacagtttc atagcctccc 780 acctatccta cacagacgaa ctcaacgttc agtgtaaagc tatagtaaag gttcacgggg 840 tctttccgtc ttgccacggg aacgctgcat cttcacagcg atttcaattt cactgagtct 900 cgggtggaga cagcgccgct gtcgttacgc cattcgtgca ggtcggaact tacccgacaa 960 ggaatttcgc taccttagga ccgttatagt tacggccgcc gtttactggg gcttcgatca 1020 agagcttcgc cttgcggctg accccatcaa ttaaccttcc agcaccgggc aggcgtcaca 1080 ccctatacgt ccactttcgt gtttgcagag tgctgtgttt ttgataaaca gtcgcagcgg 1140 cctggtttct gcgaccctct tcagctatag ctcgcatgag ccaccaaaaa gggtgcacct 1200 tctcccgaag ttacggtgcc atgttgccta gttccttcac ccgagttctc tcaagcgcct 1260 gagaattctc atcctaccca cctgtgtcgg tttacggtac ggtcttcgtg agctgaagct 1320 taggagcttt tcctggaagc gtggtatcag tgacttcgcc ataaaggctc gtctcggtgc 1380 tcggtcttaa aggatcccgg atttgccaaa gatccaaacc taccgccttt ccccgggaca 1440 accaacgccc ggtacaccta accttctccg tccctccatc gcactcacgc gaggtgcagg 1500 aatattaacc tgcttcccat cgactacggc tttcgccctc gccttaggga ccgactaacc 1560 ctgcgtcgat taacgttgcg caaggaaacc ttgggctttc ggcgtgcggg cttttcaccc 1620 gcattatcgt tactcatgtc agcattcgca cttccgatac ctccagcaga cttctcaatc 1680 caccttcgca ggcttacgga acgctcctct accgcgcata aaaccgaagt tttacgcacc 1740 ccaagcttcg gttcactgct tagccccgtt aaatcttccg cgcagaccga ctcgaccagt 1800 gagctattac gctttcttta aagggtggct gcttctaagc caacctcctg gctgtctatg 1860 cctttccaca tcgttttcca cttagcagtg aatttgggac cttagctgtg ggtctgggtt 1920 gtttcccttt tcacgacgga cgttagcacc cgccgtgtgt ctcccggata gtacgtactg 1980 gtattcggag tttgcaatgg tttggtaagt cgcgatgacc ccctagccat aacagtgctc 2040 tacccccagt agtattcgtc cgaggcgcta cctaaatagc tttcgaggag aaccagctat 2100 ctccgggttc gattagcttt tcactcctaa tcacagctca tccccgtctt ttgcaacaga 2160 cgtgggttcg ggcctccagt acctgttacg gcaccttcac cctggccatg actagatcac 2220 ccggtttcgg gtctactgcc cgcgactatg cgcccttatc agactcggtt tcccttcgcc 2280 tcccctatac ggttaagctt gccacgaaca gtaagtcgct gacccattat acaaaaggta 2340 cgcagtcact cttgcgagct cctactgctt gtacgcacac ggtttcagga tctatttcac 2400 tcccctctcc ggggttcttt tcgcctttcc ctcacggtac tggttcacta tcggtcggtc 2460 aggagtattt agccttggag gatggtcccc ccatattcag acagggtttc acgtgccccg 2520 ccctactcgt cttcactgga gtggcccttt taaatacagg gctatcacct tctatggcca 2580 atctttccag attgtttttc taaagccatt ccagcttaag ggctgttccc cgttcgctcg 2640 tcactactca gggaatctcg gttgatttct tttcctccgg ttacttagat atttcagttc 2700 accgggttcg cttcaagcag ctatgaattc actgcaagat actgccgaag cagtgggttt 2760 ccccattcgg atattgccgg atcaaagctt gttgccagct ccccgacact tttcgcaggc 2820 taccacgtcc ttcatcgcct ctgaccgcct aggcatccac cgtgtgcgct tattcgcttg 2880 acc 2883 <210> 2 <211> 2714 <212> DNA <213> X melonis BC2621 <400> 2 acacacctga cctatcaacc acgtagtcta catggttcct ttagggggct tgtgccccgg 60 gaagtctcat cttgaggcgc gcttcccgct tagatgcttt cagcggttat cgcttccgaa 120 catagctacc cggcaatgcc actggcgtga caaccggaac accagaggtt cgtccactcc 180 ggtcctctcg tactaggagc agcccctctc aaacttccaa cgcccatggc agatagggac 240 cgaactgtct cacgacgttc tgaacccagc tcgcgtacca ctttaaatgg cgaacagcca 300 tacccttggg accgactaca gccccaggat gtgatgagcc gacatcgagg tgccaaacac 360 cgccgtcgat atgaactctt gggcggtatc agcctgttat ccccggagta ccttttatcc 420 gttgagcgat ggcccttcca tacagaacca ccggatcact aagacctact ttcgtacctg 480 cttgatccgt cgatcttgca gtcaagcacg cttatgcctt tgcacacagt gcgcgatgtc 540 cgaccgcgct gagcgtacct tcgtgctcct ccgttactct ttaggaggag accgccccag 600 tcaaactacc caccatacac ggtccccgat ccggataacg gatctaggtt agaacgtcaa 660 gcacgacagg gtggtatttc aaggatggct ccactgcagc tagcgccaca gtttcatagc 720 ctcccaccta tcctacacag acgaactcaa cgttcagtgt aaagctatag taaaggttca 780 cggggtcttt ccgtcttgcc acgggaacgc tgcatcttca cagcgatttc aatttcactg 840 agtctcgggt ggagacagcg ccgctgtcgt tacgccattc gtgcaggtcg gaacttaccc 900 gacaaggaat ttcgctacct taggaccgtt atagttacgg ccgccgttta ctggggcttc 960 gatcaagagc ttcgccttgc ggctgacccc atcaattaac cttccagcac cgggcaggcg 1020 tcacacccta tacgtccact ttcgtgtttg cagagtgctg tgtttttgat aaacagtcgc 1080 agcggcctgg tttctgcgac cctcttcagc tatagctcgc acgagccacc aaaaagggtg 1140 caccttctcc cgaagttacg gtgccatgtt gcctagttcc ttcacccgag ttctctcaag 1200 cgcctgagaa ttctcatcct acccacctgt gtcggtttac ggtacggtct tcgtgagctg 1260 aagcttagga gcttttcctg gaagcgtggt atcagtgact ttgccataaa ggctcgtctc 1320 ggtgctcggt cttaaaggat cccggatttg ccaaagatcc aaacctaccg cctttccccg 1380 ggacaaccaa cgcccggtac acctaacctt ctccgtccct ccatcgcact cacgcgaggt 1440 gcaggaatat taacctgctt cccatcgact acggctttcg ccctcgcctt agggaccgac 1500 taaccctgcg tcgattaacg ttgcgcaagg aaaccttggg ctttcggcgt gcgggctttt 1560 cacccgcatt atcgttactc atgtcagcat tcgcacttcc gatacctcca gcagacttct 1620 caatccacct tcgcaggctt acggaacgct cctctaccgc gcataaaaca agttttatgc 1680 accccaagct tcggttcact gcttagcccc gttaaatctt ccgcgcagac cgactcgacc 1740 agtgagctat tacgctttct ttaaagggtg gctgcttcta agccaacctc ctggctgtct 1800 atgcctttcc acatcgtttt ccacttagca gtgaatttgg gaccttagct gtgggtctgg 1860 gttgtttccc ttttcacgac ggacgttagc acccgccgtg tgtctcccat acagtccgtc 1920 tcggtattcg gagtttgcaa tggtttggta agtcgcgatg accccctagc cataacagtg 1980 ctctaccccc gagaggatac atatgaggcg ctacctaaat agctttcgag gagaaccagc 2040 tatctccggg ttcgattagc ttttcactcc taatcacagc tcatccccgt cttttgcaac 2100 agacgtgggt tcgggcctcc agtacctgtt acggcacctt caccctggcc atgactagat 2160 cacccggttt cgggtctact gcccgcgact atgcgccctt atcagactcg gtttcccttc 2220 gcctccccta tacggttaag cttgccacga acagtaagtc gctgacccat tatacaaaag 2280 gtacgcagtc actcttgcga gctcctactg cttgtacgca cacggtttca ggatctattt 2340 cactcccctc tccggggttc ttttcgcctt tccctcacgg tactggttca ctatcggtcg 2400 gtcaggagta tttagccttg gaggatggtc cccccatatt cagacagggt ttcacgtgcc 2460 ccgccctact cgtcttcact ggagtggccc ttttaaatac agggctatca ccttctatgg 2520 ccaatctttc cagattgttt ttctaaagcc attccagctt aagggctgct ccccgttcgc 2580 tcgtcactac ttagggaatc tcggttgatt tcttttcctc cggttactta gatatttcag 2640 ttcaccgggt tcgcttccag cagctatgaa ttcactgcag gatactgccg aagcagtggg 2700 tttccccatt cgga 2714 <210> 3 <211> 27 <212> DNA <213> Xan23SF5 <400> 3 tggtcaagcc gcacggatca ttagtat 27 <210> 4 <211> 25 <212> DNA <213> Xan23SR6 <400> 4 acgtggatag cctgcgaaaa gtgtc 25 <210> 5 <211> 22 <212> DNA <213> Xan23SF7 <400> 5 gagaccgccc cagtcaaact ac 22 <210> 6 <211> 23 <212> DNA <213> Xan23SR7 <400> 6 accttttgta taatgggtca acg 23 <210> 7 <211> 25 <212> DNA <213> Xan23SF9 <400> 7 gtcaagccgc acggatcatt agtat 25 <210> 8 <211> 20 <212> DNA <213> Xan23SR9 <400> 8 gtcggggagc tggcaacaag 20

Claims (4)

(a) separating 23S rRNA from the sample;
(b) confirming the nucleotide sequence of the 23S rRNA isolated in the step (a); And
(c) the nucleotide sequence identified in step (b) is the 78th base G of SEQ ID NO: 1, the 86th base C of SEQ ID NO: 1, the 682th base C of SEQ ID NO: 1, the 740th base A of SEQ ID NO: Base 1177 of SEQ ID NO: 1, nucleotide T at position 1357 of SEQ ID NO: 1, nucleotide T of nucleotide 1735 of SEQ ID NO: 1, nucleotide 1966 of base 1966 of SEQ ID NO: 1, nucleotide 1967 of nucleotide 1967 of SEQ ID NO: 1, nucleotide 1969 The nucleotide 1973 of SEQ ID NO: 1, the nucleotide 1977 of SEQ ID NO: 1, the nucleotide 1978 of SEQ ID NO: 1, the nucleotide 1979 of SEQ ID NO: 1, the nucleotide 2048 of SEQ ID NO: 1, The 2049th base G of SEQ ID NO: 1, the 2053th base G of SEQ ID NO: 1, the 2056th base A of SEQ ID NO: 1, the 2058th base A of SEQ ID NO: 1, the 2060th base G of SEQ ID NO: Base A at position 2061 of SEQ ID NO: 1, nucleotide 2626 of base 262 of SEQ ID NO: 1, base 2649 at nucleotide 2649 of SEQ ID NO: 1, 2715th base C of SEQ ID NO: 1, and Xanthomonas melonis in the case of 2737th base G of SEQ ID NO: 1.
A polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 78th base G of SEQ ID NO: 1, a polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 86th base C of SEQ ID NO: 1, A polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 682nd nucleotide C of SEQ ID NO: 1, a polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 740th nucleotide A of SEQ ID NO: 1, A polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 1177th base C of SEQ ID NO: 1, a polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 1357th nucleotide T of SEQ ID NO: 1, A polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 1735th base T of SEQ ID NO: 1, 20-100 consecutive sequences comprising 1966th base A of SEQ ID NO: , A polynucleotide consisting of 20-100 consecutive DNA sequences comprising 1967 base T of SEQ ID NO: 1, 20-100 consecutive sequences comprising 1969th base C of SEQ ID NO: 1 A polynucleotide consisting of 20-100 consecutive DNA sequences comprising 1973 base C of SEQ ID NO: 1, 20-100 consecutive sequences comprising 1977 base C of SEQ ID NO: 1 A polynucleotide consisting of 20-100 consecutive DNA sequences comprising 1978 base T of SEQ ID NO: 1, 20-100 consecutive sequences comprising 1979 base C of SEQ ID NO: 1 A polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 2048th base G of SEQ ID NO: 1, a polynucleotide consisting of a polynucleotide consisting of a nucleotide sequence of SEQ ID NO: A polynucleotide consisting of 20-100 consecutive DNA sequences comprising 2049th base A of SEQ ID NO: 1, a polynucleotide consisting of 20-100 consecutive DNA sequences comprising 2050th nucleotide G of SEQ ID NO: 1, 1, a polynucleotide consisting of 20-100 consecutive DNA sequences containing the 2053 base G of SEQ ID NO: 1, a polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 2056th base A of SEQ ID NO: 1, A polynucleotide consisting of 20-100 consecutive DNA sequences containing the 2058th base A of SEQ ID NO: 1, a polynucleotide consisting of 20-100 consecutive DNA sequences comprising 2060th base A of SEQ ID NO: 1, 1, 20-100 consecutive DNA sequences comprising 2061th base T, 20-100 consecutive Ds comprising 2626th nucleotide T of SEQ ID NO: 1 A polynucleotide consisting of 20-100 contiguous DNA sequences comprising the 2649th base T of SEQ ID NO: 1, a polynucleotide consisting of 20-100 consecutive contiguous DNA sequences comprising 2715th C of SEQ ID NO: 1 A polynucleotide consisting of 20-100 consecutive DNA sequences comprising the 2737th base G of SEQ ID NO: 1, and Xanthomonas melonis including both complementary polynucleotides thereof. ) Identification probe composition.
A microarray for detecting Xanthomonas melonis characterized in that the probe composition of claim 2 is integrated on a substrate.
In the nucleotide having the nucleotide sequence shown in SEQ ID NO: 1, the 78th base G of SEQ ID NO: 1, the 86th nucleotide C of SEQ ID NO: 1, the 682nd nucleotide C of SEQ ID NO: 1, the 740th nucleotide A of SEQ ID NO: 1, The nucleotide sequence of nucleotides 1177 of SEQ ID NO: 1, nucleotide T of nucleotide 1357 of SEQ ID NO: 1, nucleotide 1735 of nucleotide T of SEQ ID NO: 1, nucleotide 1966 of nucleotide 1966 of SEQ ID NO: 1, nucleotide 1967 of nucleotide 1967 of SEQ ID NO: 1, nucleotide 1969 Base C of 1973, base 1977 of base sequence C of SEQ ID NO: 1, base 1978 of base sequence T of SEQ ID NO: 1, base 1979 of base 1979 of SEQ ID NO: 1, base 2048 of base sequence G of SEQ ID NO: 1, 1, 2050th base G of SEQ ID NO: 1, 2053th base G of SEQ ID NO: 1, 2056th base A of SEQ ID NO: 1, 2058th base A of SEQ ID NO: 1, 2060th base of SEQ ID NO: 1 A, nucleotide 2061 of SEQ ID NO: 1, nucleotide 2626 of SEQ ID NO: 1, nucleotide 264 of SEQ ID NO: 1 The 9th base T, the 2715th base C of SEQ ID NO: 1, and the 2737th base G of SEQ ID NO: 1.

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