Abstract

ARANA LABRADA, Franklyn et al. Development of a method for the specific diagnosis of PepMoV based on RT -PCR. Rev. Protección Veg. [online]. 2016, vol.31, n.1, pp. 20-28. ISSN 2224-4697.

The main objective was to develop a method for the specific diagnosis of PepMoV based on RT-PCR. Primers were designed using as reference the genomic sequence of the Florida isolate to amplify a fragment of the 5'untranslatable end and amino-terminus of P1 protein of the genome. The Oligo program (version 7.4) was used to carry out the analysis in silico and establish the initial parameters of the reaction. The optimal temperature of primer hybridization, primer concentration, intra- and inter-assay repeatability, detection limit, and the analytic specificity were evaluated. The optimal temperature of primer hybridization was 59°C, and 0,32 µM was selected as the optimal concentration of the primers. The aspects of the assay evaluated allowed establishing the optimal conditions of the technique. The RT-PCR method behaved with a high specificity for the diagnosis of PepMoV. It showed a detection limit of 94 pg.µl^-1. The molecular tool obtained will allow supporting the breeding programs and the integrated pest management of this crop in the country.

Keywords : RT-PCR; potyviruses; PepMoV; sweet pepper.

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