53 Identification of Simple Sequence Repeat (SSR) Markers Linked to Flowering Time in Almond by Bulked Segregant Analysis (BSA) M. Rasouli 1,2 , R. Fatahi 2 , Z. Zamani 2 , A. Imani 3 and P. Martínez-Gómez 4,a 1 Horticulture Science Department, Faculty of Agriculture, University of Tehran, Karaj, Iran 2 Department of Landscape Engineering, Faculty of Agriculture, Malayer University, Malayer, Iran 3 Department of Horticulture, Seed and Plant Improvement Institute, Karaj, Iran 4 Department of Plant Breeding, CEBAS-CSIC, P.O. Box 164, 30100 Campus Universitario, Espinardo, Murcia, Spain Keywords: almond, Prunus dulcis, breeding, cultivar, fingerprinting, flowering, molecular markers Abstract In this study, flowering dates were evaluated during two years in a F 1 almond progeny of seventy eight seedlings from the cross between the intermediate flowering Italian cultivar ‘Tuono’ and the extra-late flowering Iranian cultivar ‘Shahrood-12’. In addition, a modified-bulk segregant analysis in combination with the application of SSR markers was used to identify molecular markers linked to flowering time. Seventy five nuclear SSR markers spanning the whole almond genome were assayed in the molecular characterization of bulks (extra-early, early, late and extra-late) consisting in several descendants selected from the almond progeny studied. Results showed a quantitative inheritance of flowering date in the almond progeny studied. The seedlings evaluated showed a wide range of flowering dates between both progenitors although some of these descendants were earlier than the early progenitor ‘Tuono’. Two microsatellite loci (CPPCT008 and EPDCU2584) were found to be tightly linked to this important agronomic trait. Finally, we discuss the development of efficient marker-assisted selection strategies applied to almond and other Prunus breeding programs. INTRODUCTION Evaluation of flowering time in almond is a tedious process because of the long juvenile period of trees, the influence of the juvenility on the expression of the trait, and the existence of climatic factors affecting this evaluation. Marker-assisted selection (MAS) is particularly useful in these cases (Arús and Moreno-González, 1993). The main approach for the development of molecular markers for MAS strategies in almond has been the use of segregating progenies and the identification of quantitative trait loci (QTLs) linked to these traits (Martínez-Gómez et al., 2012). However, bulked segregant analysis (BSA), where two pooled DNA samples are formed from plant sources that have similar genetic backgrounds but differ in one particular trait, is another powerful approach for the analysis of molecular marker-horticultural trait association (Michelmore et al., 1991). In the BSA analysis, the segregating populations are evaluated phenotypically as a first step. Then, genotypic evaluation is performed on only a subset of the population: those genotypes that occur in the tails of the distribution of the trait of interest. By genotyping a subsample of the population, the cost of an association study can be significantly reduced. This selective genotyping is used when growing and phenotyping individuals in a mapping population are easier and/or cheaper than genotyping using DNA markers. On the other hand, DNA simple sequence repeats (SSR) markers are based on the PCR technique through the specific amplification of the conserved DNA sequence flanking repetitive DNA sequences (microsatellite loci) of the genome (Tauzt, 1989). a pmartinez@cebas.csic.es Proc. VI th IS on Almonds and Pistachios Eds.: F. Dicenta et al. Acta Hort. 1028, ISHS 2014