The advance of Next Generation Sequencing (NGS) technologies allows high-throughput genotyping at a reasonable cost, although, in the case of peach, this technology has been scarcely developed. To date, only a standard Genotyping by Sequencing approach (GBS), based on a single restriction with ApeKI to reduce genome complexity, has been applied in peach. In this work, we assessed the performance of the double-digest RADseq approach (ddRADseq), by testing 6 double restrictions with the restriction profile generated with ApeKI. The enzyme pair PstI/MboI retained the highest number of loci in concordance with the in silico analysis. Under this condition, the analysis of a diverse germplasm collection (191 peach genotypes) yielded 200,759,000 paired-end (2 × 250 bp) reads that allowed the identification of 113,411 SNP, 13,661 InDel and 2133 SSR. We take advantage of a wide sample set to describe technical scope of the platform. The novel platform presented here represents a useful tool for genomic-based breeding for peach.EEA San PedroFil: Aballay, Maximiliano Martín. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria San Pedro; Argentina.Fil: Aballay, Maximiliano Martín. Consejo Nacional de Investigaciones Científica y Técnicas; ArgentinaFil: Aguirre, Natalia Cristina. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina.Fil: Aguirre, Natalia Cristina. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Agrobiotecnología y Biología Molecular; Argentina.Fil: Filippi, Carla Valeria. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; ArgentinaFil: Filippi, Carla Valeria. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Agrobiotecnología y Biología Molecular; ArgentinaFil: Valentini, Gabriel Hugo. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria San Pedro; ArgentinaFil: Sánchez, Gerardo. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria San Pedro; Argentin
