Disease Resistance Gene Candidate Sequences in Gerbera: Cloning, Characterization, and Utilization for Molecular Marker Development

The most common class of disease resistance genes (R genes) in plant genomes seems to be the nucleotide-binding site leucine-rich repeat (NB-LRR) class genes. High levels of sequence conservation in this class of R genes have enabled the designing of degenerate primers for PCR amplification of resistance gene candidate (RGC) sequences from numerous plants. In this study, eight combinations of reported degenerate primers were used to amplify RGCs from gerbera. Out of 172 fragments sequenced, 84 were RGC sequences containing the typical motifs of the NB domain of the NB-LRR R genes. Twenty-eight representative gerbera RGC sequences were selected for further analysis, and they were clustered into nine groups, designated as RGC1 to RGC9, respectively. RGC1 to RGC4 belong to the TIR (Toll interleukin receptor)-NB-LRR subfamily, while RGC5 to RGC9 to the CC (coiled coil)-NB-LRR subfamily of plant NB-LRR R genes. Specific primers designed from 15 of the RGCs detected polymorphisms, following restriction enzyme digestion, between the gerbera breeding line UFGE 4033 and the gerbera cultivar ‘Sunburst Snow White’, parents of two mapping populations that were created for locating and mapping genes for powdery mildew resistance in gerbera. When used with four random primers designed for the target region amplification polymorphism marker system, RGC-derived specific primers revealed additional polymorphisms between the two parents. Our results indicate that RGC sequences can be very useful and valuable in multiple ways for molecular marker development.