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Open Access Development of a real-time PCR assay for the detection of Cladosporium fulvum in tomato leaves

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Abstract Aims: 

The aim of this study was to develop a sensitive real-time polymerase chain reaction (PCR) assay for the rapid detection of Cladosporium fulvum in tomato leaves. Methods and Results: 

Three PCR primer pairs were designed based on the nucleotide sequences of: (i) the internal transcribed spacer regions of ribosomal RNA; (ii) a microsatellite region amplified by the microsatellite primer M13; and (iii) the β-tubulin gene of C. fulvum. Each primer pair amplified the expected target DNA fragment from geographically diverse isolates of C. fulvum. No PCR products were amplified with these primer pairs from DNA of other fungal species. Among the three pairs of primers, the primer pair CfF1/CfR1 developed based on the microsatellite region was the most sensitive. Using this sensitive primer pair, a real-time PCR assay was developed to detect early infection of C. fulvum in tomato leaves. Significance and Impact of the Study: 

DNA regions amplified by the microsatellite primer M13 have a high potential for developing highly sensitive species-specific PCR primers for the detection of phytopathogenic fungi. The real-time PCR assay developed in this study is useful in monitoring early infection of C. fulvum, and can help growers make timely decisions on fungicide application.
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Keywords: Cladosporium fulvum; leaf mould of tomato; microsatellite region; real-time PCR

Document Type: Research Article

Publication date: 01 May 2008

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