Plant & Animal Genomes XIII Conference Plant & Animal Genomes XIII ConferenceJanuary 15-19, 2005
Town & Country Convention Center
San Diego, CA
Dario A. Palmieri , Gustavo Astua-Monge , Ana C. Basilio , Vinicius de Lima , Gisele Bacocina , Juliana G. T. Roncoletta , Marcos A. Machado
Citrus breeding has been traditionally hindered by different botanical and genetic factors, which, in part, have been overcome by the use of molecular markers. Microsatellites or simple sequence repeats (SSRs) have been extensively exploited for genome mapping and for a wide range of population and evolutionary studies in several plant species. The CitEST (Integration of Genetic Breeding, Functional and Comparative Genomics of Citrus) project has generated a considerable number of EST (Expressed Sequence Tags) sequences from several citrus species and relatives. These ESTs are a potential source for the discovery of SSRs markers, which would be a useful tool for both genetic mapping and germplasm characterization studies. We report on the developing of a SSR identification pipeline that integrates PERL scripts, the Primer3 program, an SQL database, PHP scripts and HTML. This pipeline was used to screen more than 100,000 citrus ESTs in order to identify potential SSR markers and assess their polymorphism in silico. More of 9,300 candidate SSRs were identified from sweet orange, mandarins and trifoliate orange. To validate the pipeline efficiency and the predicted polymorphism, primer pairs were synthesized for 16 candidate SSRs, and screened using a set of 12 DNA samples representing different citrus genotypes. Results indicated that all tested markers exhibited the polymorphism predicted by the pipeline. This high-throughput approach is now being used to massively develop SSR markers in citrus.