Development of a Multiple Detection Technique for Fungi by DNA Microarray with the Simultaneous Use of Internal Transcribed Spacer Region of Ribosomal RNA Gene and β-Tubulin Gene Probes
We offer the first description of the development of a multiple detection technique for fungi by DNA microarray with the simultaneous use of internal transcribed spacer region (ITS) of ribosomal RNA gene and β-tubulin gene probes. The assay uses 12 oligonucleotide probes and multiplex amplification...
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| Veröffentlicht in: | Biocontrol Science 2014, Vol.19(3), pp.139-145 |
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| creator | ISSHIKI, ATSUNORI TAKEHARU, HITOSHI AOKI, SHUNSUKE KOKAJI, MAMI TANABE, SUGURU KASETANI, TAISUKE YOSHIDA, MITSUHIRO |
| description | We offer the first description of the development of a multiple detection technique for fungi by DNA microarray with the simultaneous use of internal transcribed spacer region (ITS) of ribosomal RNA gene and β-tubulin gene probes. The assay uses 12 oligonucleotide probes and multiplex amplification to detect fungal species belonging to various sections of Aspergillus, the Eurotium genus, and the Penicillium genus. The specificity of each probe was tested using 231 reference fungal strains, including 79 target and 152 non-target strains in 102 species of 24 genera. We determined the optimum concentration of the primer pairs for multiplex PCR to be 0.5 μM for the β-tubulin gene and 0.125 μM for the ITS region. In the field trial using 76 specimens containing 323 fungi (up to five fungal strains were included in one specimen), the concordance rate between the DNA microarray and the DNA sequencing results was 97.4％ at the species or genus levels. |
| doi_str_mv | 10.4265/bio.19.139 |
| format | Article |
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The assay uses 12 oligonucleotide probes and multiplex amplification to detect fungal species belonging to various sections of Aspergillus, the Eurotium genus, and the Penicillium genus. The specificity of each probe was tested using 231 reference fungal strains, including 79 target and 152 non-target strains in 102 species of 24 genera. We determined the optimum concentration of the primer pairs for multiplex PCR to be 0.5 μM for the β-tubulin gene and 0.125 μM for the ITS region. In the field trial using 76 specimens containing 323 fungi (up to five fungal strains were included in one specimen), the concordance rate between the DNA microarray and the DNA sequencing results was 97.4％ at the species or genus levels.</description><subject>Aspergillus - genetics</subject><subject>Aspergillus - isolation & purification</subject><subject>DNA microarray</subject><subject>DNA, Fungal - genetics</subject><subject>DNA, Ribosomal Spacer - genetics</subject><subject>Eurotium - genetics</subject><subject>Eurotium - isolation & purification</subject><subject>Fungi</subject><subject>ITS region</subject><subject>Microbiological Techniques - methods</subject><subject>Molecular Diagnostic Techniques - methods</subject><subject>Multiplex detection</subject><subject>Oligonucleotide Array Sequence Analysis - methods</subject><subject>Oligonucleotide Probes - genetics</subject><subject>Penicillium - genetics</subject><subject>Penicillium - isolation & purification</subject><subject>Sensitivity and Specificity</subject><subject>Tubulin - genetics</subject><subject>β-tubulin gene</subject><issn>1342-4815</issn><issn>1884-0205</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2014</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNo9kc9q3DAQh0Vpaf60lz5AmXPBG8mSvPahh5Bt0kDSls3mbCR7vKtgS64kN-xr5ZiH6DNFyyaLQBLDxzfM_Aj5wuhM5IU808bNWDVjvHpHjllZiozmVL5Pfy7yTJRMHpGTEB4oLcqykh_JUS7TKURxTJ4X-A97Nw5oI7gOFNxOfTRjj7DAiE00zsIKm401fyeEznm4nOzagN7C4tc53JrGO-W92sKjiRuIG4Q7MySHsuimAPcBd95rG9Fb1cPKKxsabzS2cDeqBj0scb3rkqil0S64IWHL5L5Ci6BsC_-fstWkp97Yfe2PdxrDJ_KhU33Az6_vKbm__LG6-Jnd_L66vji_yRpJRcwazUqGTEoqq7QBwVDzqmVUqfmcdyzdBedlMde0E7Ls2kJwngssRde2ncyRn5Jve2-aNASPXT16Myi_rRmtdwHUKYCaVXUKIMFf9_A46QHbA_q28QR83wMPIao1HgDlo2l6fHPxV-Gh3myUr9HyF7tvmho</recordid><startdate>2014</startdate><enddate>2014</enddate><creator>ISSHIKI, ATSUNORI</creator><creator>TAKEHARU, HITOSHI</creator><creator>AOKI, SHUNSUKE</creator><creator>KOKAJI, MAMI</creator><creator>TANABE, SUGURU</creator><creator>KASETANI, TAISUKE</creator><creator>YOSHIDA, MITSUHIRO</creator><general>The Society for Antibacterial and Antifungal Agents, Japan</general><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope></search><sort><creationdate>2014</creationdate><title>Development of a Multiple Detection Technique for Fungi by DNA Microarray with the Simultaneous Use of Internal Transcribed Spacer Region of Ribosomal RNA Gene and β-Tubulin Gene Probes</title><author>ISSHIKI, ATSUNORI ; 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| source | J-STAGE Free; MEDLINE; Elektronische Zeitschriftenbibliothek - Frei zugängliche E-Journals |
| subjects | Aspergillus - genetics Aspergillus - isolation & purification DNA microarray DNA, Fungal - genetics DNA, Ribosomal Spacer - genetics Eurotium - genetics Eurotium - isolation & purification Fungi ITS region Microbiological Techniques - methods Molecular Diagnostic Techniques - methods Multiplex detection Oligonucleotide Array Sequence Analysis - methods Oligonucleotide Probes - genetics Penicillium - genetics Penicillium - isolation & purification Sensitivity and Specificity Tubulin - genetics β-tubulin gene |
| title | Development of a Multiple Detection Technique for Fungi by DNA Microarray with the Simultaneous Use of Internal Transcribed Spacer Region of Ribosomal RNA Gene and β-Tubulin Gene Probes |
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