Submitted to: Phytopathology
Publication Type: Abstract Only
Publication Acceptance Date: 5/22/1998
Publication Date: N/A
Technical Abstract: A simple and sensitive assay was developed for detection of potato ring rot bacterium, Clavibacter michiganensis sepedonicus (C.m.s.) in seed potatoes and stem tissues. The bacterium DNA was extracted from 2 g of chopped tuber or stem tissue using a modified alkaline DNA extraction method (S. Zhang and P.H. Goodwin. 1997. J. Phytopathology 145:267-270). A C.m.s.-specific DNA sequence was amplified in the polymerase chain reaction (PCR) primed by digoxigenin- (Dig-) labeled primer pair CMSIF1/CMSIR1 (or CMSIF2/CMSIR2), which was previously designed based on insertion element IS1121 of (C.m.s.) Dig-labeled PCR products were then blotted on a nylon membrane and the signal was detected using a colorimetric alkaline phosphatase substrate. No signal was detected from the PCR control devoid of target DNA. Signals indicating the presence of C.m.s. were observed using extracts from symptomatic potatoes and from a fraction of symptomless commercial potato tubers. The new assay procedure was equivalent to nested PCR in sensitivity of C.m.s. detection and was as simple to use as the standard ELISA procedure that is used commerically.