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Evaluation of Primer Pairs for the Reliable Diagnosisof Paulownia Witches'-Broom Disease Using a Polymerase Chain Reaction. H. Nakamura, Faculty of Agriculture, Iwate University, Ueda 3-chome, Morioka 020, Japan. N. Yoshikawa, and T. Takahashi, Faculty of Agriculture, Iwate University, Ueda 3-chome, Morioka 020, Japan; N. Sahashi, T. Kubono, and T. Shoji, Tohoku Research Center, Forestry and Forest Products Research Institute, Shimokuriyagawa, Morioka 020-01, Japan. Plant Dis 80:302. Accepted for publication 20 November 1995. Copyright 1996 The American Phytopathological Society. DOI: 10.1094/PD-80-0302.

To establish a reliable method for detection of paulownia witches'-broom (PWB) phytoplasma by a polymerase chain reaction (PCR), several oligonucleotide primers that amplify ribosomal protein (rp) or 16S rRNA (rD) genes of PWB-phytoplasma were compared for their specificity and sensitivity in the amplification of PWB-specific fragments by PCR. Use of rp primer pairs for PCR resulted in amplified DNA fragments of expected sizes in samples from diseased leaves. A fragment was amplified from healthy samples using one primer pair (rp3/rp4), but the fragment size was different from disease-specific fragments. In contrast, all four rD primer pairs designed for the 16S rRNA gene amplified both PWB-specific DNA fragments from infected leaf samples and nonspecific fragments of the same size as PWB-specific fragments from healthy samples. Using the rp3/rp4 primer pair, we amplified a PWB-specific DNA fragment from 150 pg of nucleic acid samples. This method allowed detection at the 95% confidence level of PWB-phytoplasma from paulownia leaves from the ten asymptomatic infected trees used in this study when at least 3 leaves randomly collected in September were used.