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Nucleic Acid Probes for Identification of Phytobacteria: Identification of Genus-Specific 16s rRNA Sequences. J. DeParasis, Department of Plant, Soil and Insect Sciences, University of Wyoming, Laramie 82071; Don A. Roth, Department of Plant, Soil and Insect Sciences, University of Wyoming, Laramie 82071. Phytopathology 80:618-621. Accepted for publication 3 January 1990. Copyright 1990 The American Phytopathological Society. DOI: 10.1094/Phyto-80-618.

The development of DNA probes for the identification of phytobacteria requires efficient procedures to identify target-specific nucleic acid sequences present in high copy number within the target cells. Probes based on phytobacterial rRNA sequences should provide an extremely sensitive identification method. To establish the potential specificity of this approach, 16s rRNAs of 52 strains were partially sequenced. The approach was based on partially purifying high molecular weight RNAs from crude lysates and selecting the 16s rRNA by hybridization with oligodeoxynucleotide primers to conserved regions. Reverse transcription and dideoxynucleotide chain termination methods allowed the sequencing of 600?800 nucleotides of the approximate total 1,540 comprising 16s rRNA. Although most of the molecule is highly conserved, a region defined by the bases Nos. 1057?1090 (numbered from the cDNA) is variable between the genera assayed. Pathovars or subspecies could not be differentiated based on comparisons within this region. The degree of variation in the conserved molecule is sufficiently high to expect specific discrimination between phytobacterial genera using a synthetic oligodeoxynucleotide probe.