Rasmy,, M. (2002). EVALUATION OF THE POLYMERASE CHAIN REACTION (BIO-PCR) , ELISA AND SEMISELECTIVE MEDIUM FOR DETECTION TOMATO BACTERIAL CANKER.. Journal of Plant Production, 27(5), 2935-2944. doi: 10.21608/jpp.2002.254183
M. R. Rasmy,. "EVALUATION OF THE POLYMERASE CHAIN REACTION (BIO-PCR) , ELISA AND SEMISELECTIVE MEDIUM FOR DETECTION TOMATO BACTERIAL CANKER.". Journal of Plant Production, 27, 5, 2002, 2935-2944. doi: 10.21608/jpp.2002.254183
Rasmy,, M. (2002). 'EVALUATION OF THE POLYMERASE CHAIN REACTION (BIO-PCR) , ELISA AND SEMISELECTIVE MEDIUM FOR DETECTION TOMATO BACTERIAL CANKER.', Journal of Plant Production, 27(5), pp. 2935-2944. doi: 10.21608/jpp.2002.254183
Rasmy,, M. EVALUATION OF THE POLYMERASE CHAIN REACTION (BIO-PCR) , ELISA AND SEMISELECTIVE MEDIUM FOR DETECTION TOMATO BACTERIAL CANKER.. Journal of Plant Production, 2002; 27(5): 2935-2944. doi: 10.21608/jpp.2002.254183
EVALUATION OF THE POLYMERASE CHAIN REACTION (BIO-PCR) , ELISA AND SEMISELECTIVE MEDIUM FOR DETECTION TOMATO BACTERIAL CANKER.
Plant Pathology Research Institute; ARC,Giza, Egypt.
Abstract
Current seed health testing is based on standardized methods described by International Seed Testing Association (ISTA). Some of these tests are time consuming. Most techniques being used for detecting seedborne bacteria utilize semiselective agar media for detecting the pathogen followed by pathogenicity tests for identification. These techniques can be very sensitive but depend upon the selectivity of the medium and contamination level of the samples with saprophytes. Semiselective agar media have a sensitivity range between 1x10¹ to 1 x 10² cfu/ml. This is 100-1,000 fold more sensitive than ELISA. Serological techniques (ELISA) have the advantage of producing quicker results but have the disadvantage of a relatively low detection threshold around 1 x 105 cfu/ml. Semiselective agar media have several disadvantage, including overgrowth of the target bacterium by saprophytic bacteria contaminated the seed, antibiotics produced by saprophytes, and time consuming. Molecular diagnostics offers the potential for sensitive and specific detection of Clavibacter michiganensis subsp. michiganensis (CMM). Advantages of BIO-PCR, over the existing PCRtechniques include the elimination of false positives results due to the presence of dead cells that may be present on the tomato seeds, elimination of false negatives results due to potential PCR inhibitors in seed extracts and increased sensitivity of detection.. The BIO- PCR technique is specific for CMM without amplification the closely related bacterium Clavibacter michiganensis subsp. sepedonicus, or other bacterial species. The assay was successfully applied to the detection of CMM in tomato seeds and was sensitive to approximately 10 cells per PCR assay. The described PCR method for identification of this pathogen is very fast (1-2 days) and economical, due to the very small volume (10 µl.) of the PCR reaction mixture. this method is based on amplification of the bacterial plasmid DNA fragment and may be very useful in routine identification of CMM and could be an alternative tool for diagnoses, especially for quarantine work..
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