Development of a loop-mediated isothermal amplification assay for detection of Phytophthora sojae

Phytophthora sojae is a devastating pathogen that causes soybean Phytophthora root rot. This study reports the development of a loop-mediated isothermal amplification (LAMP) assay targeting the A3aPro element for visual detection of P.similar to sojae. The A3aPro-LAMP assay efficiently amplified the target element in <similar to 80 similar to min at 64 similar to degrees C and was evaluated for specificity and sensitivity. The specificity was evaluated against P.similar to sojae, Phytophthora spp., Pythium spp., and true fungi isolates. Magnesium pyrophosphate resulting from the LAMP of P.similar to sojae could be detected by real-time measurement of turbidity. Phytophthora sojae DNA products were visualized as a ladder-like banding pattern on 2% gel electrophoresis. A positive colour (sky blue) was only observed in the presence of P.similar to sojae with the addition of hydroxynaphthol blue prior to amplification, whereas none of other isolates showed a colour change. The detection limit of the A3aPro-specific LAMP assay for P.similar to sojae was 10 similar to pg similar to mu L-1 of genomic DNA per reaction. The assay also detected P.similar to sojae from diseased soybean tissues and residues. These results suggest that the A3aPro-LAMP assay reported here can be used for the visual detection of P.similar to sojae in plants and production fields.