caa a22 4500 445390514 CHVBK 20180317143045.0 cr unu---uuuuu 170323e20110501xx s 000 0 eng 10.1007/s10327-011-0305-2 doi (NATIONALLICENCE)springer-10.1007/s10327-011-0305-2 In silico genomic subtraction guides development of highly accurate, DNA-based diagnostics for Ralstonia solanacearum race 3 biovar 2 and blood disease bacterium [Elektronische Daten] [Ryo Kubota, Mark Schell, Gabriel Peckham, Joanne Rue, Anne Alvarez, Caitilyn Allen, Daniel Jenkins] New rapid diagnostic methods are urgently needed to discriminate the quarantine pathogen Ralstonia solanacearum (Rs) race 3 biovar 2 (R3B2) from other populations of Rs that lack the adaptation to cause bacterial wilt disease in temperate regions. We used an in silico bioinformatic approach to identify several genome sequences potentially specific to R3B2 strains. Primer sets were designed to PCR-amplify sequences in these regions, and four sets were ultimately shown to be >99% accurate for detection of R3B2 strains. On the basis of these results, several primers were designed to enable development of a loop-mediated isothermal amplification assay that was rapid, technologically simple, and essentially 100% accurate for identification of R3B2 when applied to a comprehensive collection of geographically diverse Rs strains. We fortuitously found that a sequence in one of the "R3B2-specific” regions has ~90% identity to a sequence present in strains of the blood disease bacterium (BDB), a member of the Rs species complex that infects banana. Alignments of these sequences allowed design of a second PCR primer set that proved 100% accurate for identification of BDB strains when tested on the 22 BDB strains available to us. These results demonstrate the power of in silico genomic subtraction for rapid identification of population-specific DNA sequences and for the development of simple, reliable detection methods for Rs subpopulations. The Phytopathological Society of Japan and Springer, 2011 Loop-mediated isothermal amplification nationallicence Bacterial wilt nationallicence Potato brown rot nationallicence PCR nationallicence Kubota Ryo Department of Molecular Biosciences and Bioengineering, University of Hawaii, 96822, Honolulu, USA aut Schell Mark Departments of Microbiology and Plant Pathology, University of Georgia, 30602, Athens, USA aut Peckham Gabriel Department of Plant and Environmental Protection Sciences, University of Hawaii, 96822, Honolulu, USA aut Rue Joanne Departments of Microbiology and Plant Pathology, University of Georgia, 30602, Athens, USA aut Alvarez Anne Department of Plant and Environmental Protection Sciences, University of Hawaii, 96822, Honolulu, USA aut Allen Caitilyn Department of Plant Pathology, University of Wisconsin, 53706, Madison, USA aut Jenkins Daniel Department of Molecular Biosciences and Bioengineering, University of Hawaii, 96822, Honolulu, USA aut Journal of General Plant Pathology Springer Japan 77/3(2011-05-01), 182-193 1345-2630 77:3<182 2011 77 10327 https://doi.org/10.1007/s10327-011-0305-2 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1007/s10327-011-0305-2 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Kubota Ryo Department of Molecular Biosciences and Bioengineering, University of Hawaii, 96822, Honolulu, USA aut NATIONALLICENCE 700 1- Schell Mark Departments of Microbiology and Plant Pathology, University of Georgia, 30602, Athens, USA aut NATIONALLICENCE 700 1- Peckham Gabriel Department of Plant and Environmental Protection Sciences, University of Hawaii, 96822, Honolulu, USA aut NATIONALLICENCE 700 1- Rue Joanne Departments of Microbiology and Plant Pathology, University of Georgia, 30602, Athens, USA aut NATIONALLICENCE 700 1- Alvarez Anne Department of Plant and Environmental Protection Sciences, University of Hawaii, 96822, Honolulu, USA aut NATIONALLICENCE 700 1- Allen Caitilyn Department of Plant Pathology, University of Wisconsin, 53706, Madison, USA aut NATIONALLICENCE 700 1- Jenkins Daniel Department of Molecular Biosciences and Bioengineering, University of Hawaii, 96822, Honolulu, USA aut NATIONALLICENCE 773 0- Journal of General Plant Pathology Springer Japan 77/3(2011-05-01), 182-193 1345-2630 77:3<182 2011 77 10327 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer