Euphytica 129: 33–41, 2002. © 2002 Kluwer Academic Publishers. Printed in the Netherlands. 33 Identification of olive-tree cultivars with SCAR markers Roc´ ıo Bautista, Remedios Crespillo, Francisco M. C´ anovas & M. Gonzalo Claros ∗ Departamento de Biolog´ ıa Molecular y Bioqu´ ımica, Facultad de Ciencias e Instituto Andaluz de Biotecnolog´ ıa, Unidad Asociada UMA-CSIC, Universidad de M´ alaga, M´ alaga, Spain; ( ∗ author for correspondence, e-mail: claros@uma.es) Received 17 December 2001; accepted 31 May 2002 Key words: molecular markers, Olea europaea, olive tree, SCAR Summary There is an urgent need for the development of early identification techniques in olive-trees due to the economic importance of cultivar identification in periods of expansion like now. We have been able to identify 22 olive- tree cultivars using only 10 different, specific, repeatable markers. These markers were designed by the cloning of significant RAPD bands obtained in PCR performed on bulked DNA to retain the genetic variability of each cultivar. Clones were partially or totally sequenced and new primers derived from these sequences were used to obtain Sequence Characterised Amplified Region (SCAR) fragments. We have demonstrated that the use of the 10 SCAR markers is enough to provide a simple, cheap, and reliable procedure to identify 22 geographically related olive-tree cultivars. Abbreviations: AFLP – amplification fragment length polymorphism; AP-PCR – arbitrarily primed PCR; ASAP – allele-specific associated primers; CAPS – cleaved amplified polymorphic sequence; CTAB – N-cetyl-N,N,N- trimethyl-ammonium bromide; PCR – polymerase chain reaction; RAPD – random-amplified polymorphic DNA; RFLP – restriction fragment length polymorphism; SCAR – sequence-characterised amplified region; Ta – annealing temperature; UPGMA – unweighted pair group method with arithmetic averages Introduction Accurate and rapid identification of clones, variet- ies, or species is especially important in vegetatively propagated plants like olive-trees (Olea europaea L.). Hundreds of cultivars have been selected over the cen- turies for their adaptation to microclimates and soil types. As a result, there is considerable uncertainty about the names of many olive cultivars, becoming synonymous and homonymous a long-standing prob- lem in olive-producing countries. This is an unliked scenario since olive-tree cultivar identification is of great economic importance in periods of expansion in its cultivation, like now. Differentiation among olive- tree cultivars is traditionally supported by numerous phenotypic traits in maturity for trunk, leaf, flower and fruit shape. More recently, workers have found al- lozymes to be useful markers for an objective cultivar identification. The weakness of both classifications has been put in evidence by the demonstration that chemical and morphological changes in olive-trees as well as other plants are induced by domestication, among other things. For example, agronomic selec- tion for increased yield and growth rate has caused a reduction in plant defences (Massei & Hartley, 2000). Additionally, the long juvenile stage and the long non-productive period hamper a traditional method to identify the cultivar, which leads to an increase of plantation costs mainly in cases of cultivar miss- identification. Therefore, there is an urgent need for the development of methods that identify cultivars easily, rapidly and at an early stage of development. Olive cultivar identification has been rapidly im- proved using molecular markers based on DNA since they provide an opportunity for direct comparison and identification of different genetic material independent