Characterization and detection of Phytophthora fragariae in plant, water and soil by molecular methods
Bonants. P. J. M Gent-Pelzer. M. P. E. V Weerdt. M. H
Eppo Bulletin ; 2008 [Vol.30] Pages:525-531
Abstract
Phytophthora fragariae var. fragariae is pathogenic to strawberry only and is an EPPO A2 quarantine pest. Seventy-five isolates of P. fragariae from all over the world have been characterized by amplified fragment length polymorphism (AFLP) analysis. However, a sensitive, specific and fast method of detection is now needed, and polymerase chain reaction (PCR)-based methods have now been set up. Based upon sequences of the ITS (internal transcribed spacer) regions of rDNA of several Phytophthora species, PCR primers specific for P. fragariae have been developed together with SCRI (Dundee, GB). These primers were tested on DNA extracted from water, soil and plant material. A nested PCR procedure has been included in the currently used bait test to improve sensitivity and reliability. Also several methods have been studied to detect the amplicon of the PCR reaction: gel electrophoresis, DIAPOPS (detection of immobilized amplified products in a one-phase system), PCR-ELIS A, TaqMan and molecular beacon. With the last two methods, using the ABI 7700 detection system, quantification of target DNA is possible.
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