Molecular characterization of genetic variability and structure of olive (Olea europaea L.) germplasm collection analyzed by agromorphological traits and microsatellite markers

Turkish Journal of Agriculture and Forestry http://journals.tubitak.gov.tr/agriculture/ Research Article Turk J Agric For (2016) 40: 583-596 © TÜBİTAK doi:10.3906/tar-1602-27 Molecular characterization of genetic variability and structure of olive (Olea europaea L.) germplasm collection analyzed by agromorphological traits and microsatellite markers 1, 1 2 Karim SORKHEH *, Esmaeil KHALEGHI Department of Agronomy and Plant Breeding, Faculty of Agriculture, Shahid Chamran University of Ahvaz, Ahvaz, Iran 2 Department of Horticulture, Faculty of Agriculture, Shahid Chamran University of Ahvaz, Ahvaz, Iran Received: 06.02.2016 Accepted/Published Online: 28.03.2016 Final Version: 14.06.2016 Abstract: We studied 200 trees belonging to 20 accessions of cultivated olive (O. europaea L.) from 4 regions of origin, evaluated by means of agromorphological traits and simple sequence repeat (SSR) markers. The agromorphological traits showed high variation between genotypes and significant correlation coefficients were obtained among the values recorded in two consecutive years, 2013 and 2014. The maximum coefficient of variation for the quantitative agronomic traits was observed in fruit weight wet (13.45%), while the lowest was found in stone width (3.18%). Fruit shape index, leaf length, leaf width, and lenticel size also showed variability. With both DNA-based and agromorphological descriptors, higher levels of variability were found. Genetic variation observed among the olive germplasm at the DNA level was higher than that of the agromorphological traits, indicating the efficiency of SSR markers for detecting genetic diversity among olive genotypes and their relationships. The lack of consistency between the relationship studies performed with molecular and morphological markers could indicate that each marker system measures different aspects of olive genetic variability. Molecular data obtained by SSR markers together with morphological and agronomical characterization of olive trees confirmed the high diversity and their potential use for olive breeding. Key words: Genetic variability, microsatellite markers, Olea europaea L., STRUCTURE 1. Introduction Olive (Olea europaea subsp. europaea var. europaea) is one of the oldest agricultural tree crops in the Mediterranean basin with remarkable cultural and economic importance. The richness of the cultivated olive germplasm is an unusual case among horticultural crops, as a consequence of tree longevity and lack of turnover with new breeding genotypes (Barranco et al., 2005; Bartolini et al., 2005; Baldoni and Belaj, 2009). In spite of the richness of cultivated germplasm, olive cultivars exhibit lower genetic diversity than their wild relatives (Lumaret et al., 2004; Breton et al., 2006; Belaj et al., 2010), indicating that the latter could enrich the genetic basis of cultivated material. To date, most work has concentrated on evaluating the distribution of variability between cultivated and wild olives (Baldoni et al., 2006; Breton et al., 2006; Belaj et al., 2007; Erre et al., 2010) and on establishing the genetic relationships among the different O. europaea subspecies that are distributed beyond the Mediterranean area (Besnard et al., 2007; García-Verdugo et al., 2010). Due to recent advances in DNA technologies most of these studies * Correspondence: have been performed by means of molecular markers, simple sequence repeats (SSRs) being the most widely used. However, in spite of the drawbacks of traditional morphological description, such as environmental influences, and the need for extensive observations of mature plants, the joint use of both morphoagronomic traits and SSR markers could give the opportunity to exploit the complementary natures of these two methods (Karp et al., 1997) in evaluating the genetic diversity of wild olive trees. It is largely accepted that olive cultivar discrimination based on morphological descriptions is not completely reliable (Belaj et al., 2002, 2007, 2010, 2011); therefore, DNA molecular markers, and particularly microsatellites (SSRs), are today widely used (Bracci et al., 2011; Noormohammadi et al., 2014) to complement morphological analyses and to unambiguously identify the accessions held in collections. Genetic variation has been reported among naturally occurring olive clones in the literature with molecular markers. Clones were identified with RAPD and ISSR (Gemas et al., 2004; Gomes et al., 2008; Martins-Lopes 583 SORKHEH and KHALEGHI / Turk J Agric For 2. Materials and methods 2.1. Plant material We studied 200 trees belonging to 20 accessions of cultivated olive (O. europaea L.) from 4 regions of origin: Abosatl (Syria), T5 (unknown), Dezfoli (Iran), Mishen (USA), Masabei (Syria), Konservolia (Europe), Kaeisei (Europe), Kaylit (Europe), T21 (Europe), Khoseari (Syria), Zard (Iran), Roghani (Iran), T7 (Europe), Manzanila (Europe), Kavi (Syria), T2 (Europe), Balidi (Syria), Mari (Syria), Foji (Europe), and Koroneiki (Europe), with 12 individuals representing each region. All samples were collected from young orchards composed of olive trees 5–10 years old (Table 1). 2.2. Morphoagronomic characterization Field expeditions were carried out in autumn 2013 and 2014 in Ahvaz (south of Iran). Morphological characterization was based on olive descriptors developed by the International Olive Council (Mulas, 1999; Barranco et al., 2009), with AFLP (Strikic et al., 2010), and with microsatellites (Lopes et al., 2004; Muzzalupo et al., 2010; Albertini et al., 2011; Zaher et al., 2011; Ipek et al., 2012; Marra et al., 2013; Caruso et al., 2014; Noormohammadi et al., 2014; Abdessemed et al., 2015). Although currently there is intense research to develop reliable techniques for detecting mutations in genes, clone identification is still predominantly based on the study of phenotypic traits, integrated with molecular analyses. The present work reports the employment of morphoagronomic traits and SSR markers to investigate genetic diversity and relationships among 20 olive cultivars from different regions with high olive oil production in the world. Comparisons among these approaches were also made to assess their efficiency in evaluating genetic diversity levels and relationships among the olive trees under study. To the best of the authors’ knowledge this is the first morphoagronomic and microsatellite-based study on collecting olive cultivars. Table 1. Cultivar name, geographic origin, and use of fruits. 584 No. Cultivar Origin Kind of use 1 Zard North Iran Double use 2 Roghani North Iran Oil 3 Dezfoli North Iran Double use 4 T5 Unknown Table olive 5 Balidi Syria Table olive 6 Kavi Syria Table olive 7 T2 Europe Table olive 8 Abosatl Syria Double use 9 Mari Syria Double use 10 Khoseari Syria Table olive 11 Masabei Syria Double use 12 Mishen USA Table olive 13 Kaylit Europe Table olive 14 T21 Europe Table olive 15 C (...truncated)