Plant Protection Department, Slovenian Institute of Hop Research and Brewing, Cesta Žalskega tabora 2, SI-3310 Žalec, Slovenia
Centre for Plant Biotechnology and Breeding, Agronomy Department, Biotechnical Faculty, Jamnikarjeva 101, Ljubljana 1000, Slovenia
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Accepted for publication 2 June 2004.
Rapid polymerase chain reaction (PCR) assays were developed for the identification and detection of Verticillium albo-atrum hop pathotypes PG1 and PG2 from Slovenia. Of 17 pathotype-linked amplified fragment length polymorphism (AFLP) markers, 11 were cloned successfully and sequenced. To convert polymorphic AFLP markers into pathotype-specific sequence-characterized amplified region (SCAR) markers, 22 PG2- and 10 PG1-specific primer pairs were designed from 16 sequences. When primer specificity was tested on a wide range of Verticillium isolates, 10 PG2- and 6 PG1-specific primer pairs retained amplification specificity for V. albo-atrum Slovene hop isolates, but also amplified sequences in V. albo-atrum and V. dahliae hop isolates from different hop production areas in Europe, as well as in some isolates from other hosts. Primer combinations obtained from the AFLP-9-1 marker were specific only for V. albo-atrum PG2 isolates. The highly specific primers were used in multiplex PCR and a nested PCR to detect the V. albo-atrum PG2 pathotype in xylem tissue of hop plants. These new SCAR markers provide a valuable tool for rapid identification of V. albo-atrum PG1 and PG2 hop pathotypes.
© 2004 The American Phytopathological Society