Developing EST-SSR Markers for Gerbera Breeding and Genetic Analysis

SSR (simple sequence repeat) markers have become a very valuable tool for plant breeding and genetic research and are widely used in cultivar identification, pedigree tracing, homozogosity evaluation, genetic relatedness analysis, genetic diversity assessment, germplasm differentiation and preservation, gene mapping and tagging, marker-assisted selection, etc.  So far, no SSR markers have been reported for gerbera, a very important cut and pot flower in the United States of America and in the World.  In this study, we mined the publicly available gerbera expressed sequence tags (ESTs) in the Genbank database, identified numerous SSR-containing sequences, developed a large set of EST-SSR markers, and tested these markers in a number of breeding and genetic applications.  Of the 1,187 sequences identified out of 16,994 gerbera ESTs, 31.0% contain dinucleotide repeats, 27.9% trinucleotide, 4.8% tetranucleotide, 14.0% pentanucleotide, and 19.3% hexanucleotide.  AG (24.0%) and AAT (8.8%) motifs were the most abundant repeat types.  A total of 124 SSR primer pairs were designed based on the available gerbera ESTs.   When tested on the gerbera cultivar ‘Jaguar Tangerine’, 108 (87.1%) of them amplified DNA fragments of the expected sizes.  These primers were further tested on 47 gerbera cultivars and breeding lines, and 84 (77.8%) of them showed polymorphisms.  Sixty-six SSR markers were further tested for cultivar identification, genetic relatedness analysis, and it was found that as few as three SSR markers could differentiate all cultivars used in the study, 13 cultivars possess specific alleles, and cultivars tested could be grouped into several major clusters.