Development of a real-time fluorescence loop-mediated isothermal amplification assay for rapid and quantitative detection of Fusarium mangiferae associated with mango malformation.
Pu JinJi, Xie YiXian, Zhang He, Zhang Xin, Qi YanXiang, Peng Jun
Author Affiliation: Ministry of Agriculture Key Laboratory of Integrated Pest Management on Tropical Crops, Environmental and Plant Protection Institute, Chinese Academy of Tropical Agricultural Sciences, Haikou 571101, China.
PMPP Physiological and Molecular Plant Pathology 86 : 81-88
Abstract : Fusarium mangiferae is a major causal agent of mango malformation disease (MMD) worldwide. Rapid and accurate detection of the causal pathogen is the cornerstone of integrated disease management. In this paper, a real-time fluorescence loop-mediated isothermal amplification assay (RealAmp) was developed for quantitative detection of F. mangiferae in China. The LAMP primer set was designed based on a RAPD marker sequence and positive products were amplified only from F. mangiferae isolates, but not from any other species tested, showing a high specificity of the primer sets. The detection limit was approximately 2.26×10-4 ng/µl plasmid DNA when mixed with extracted mango DNA. Quantification of the pathogen DNA of MMD in naturally collected samples was no significant difference compared to classic real-time PCR Additionally, RealAmp assay was visual with an improved closed-tube visual detection system making the assay more convenient in diagnostics.