Application of the PCR technique to detect Phytophthora infestans in potato tubers and leaves.
A characterized repetitive sequence from Phytophthora infestans (P. infestans) was used to perform a PCR with the DNA from the four races 1, 3, 4, and 1-11. To obtain amplifiable DNA, all extractions had to be purified by DNA adsorbing spin columns. Only two out of four tested primers were well suited and gave an DNA amplificate of the same size for all four races. After optimization the detection limit of the PCR corresponded to 100 ng of freeze-dried mycelia per ml, specificity was established when testing a collection of the most important potato pathogenic fungi and bacteria. Using P. infestans zoospores to infect tuber slices, the detection threshold was determined to be two days post infection when 3 to 6 zoospores were applied. After infiltrating the tenfold concentration into potato leaves a visible PCR signal was obtained one day post infection. Further improvements of the sensitivity threshold in detecting P. infestans for breeding and prognosis purposes are discussed.
Niepold
F
F
Biologische Bundesanstalt für Land- und Forstwirtschaft, Institut für Pflanzenschutz in Ackerbau und Grünland, Braunschweig.
Schöber-Butin
B
B
D001483:Base Sequence; D017931:DNA Primers / Q000235:genetics; D004271:DNA, Fungal / Q000032:analysis / Q000235:genetics; D008969:Molecular Sequence Data; D010838:Phytophthora / Q000235:genetics / Q000302:isolation & purification*; D018515:Plant Leaves / Q000382:microbiology; D018517:Plant Roots / Q000382:microbiology; D016133:Polymerase Chain Reaction / Q000379:methods*; D012091:Repetitive Sequences, Nucleic Acid / Q000235:genetics; D012680:Sensitivity and Specificity; D011198:Solanum tuberosum / Q000382:microbiology*
D017931:DNA Primers; D004271:DNA, Fungal