Characterization of Resistance Gene Analogs in Musa acuminata Cultivars Contrasting in Resistance to Biotic Stresses

The majority of commercial banana cultivars (Musa sp.) have evolved via asexual vegetative propagation, with diversity dependent upon somatic mutation. Restricted variation has resulted in a crops with little resistance to pests and disease, and conventional breeding efforts are limited due to limited viable seed production. Numerous disease resistance genes (R-genes / R-proteins) have been characterized in plants, recognizing and conferring resistance to bacteria, virus, fungi and nematodes. The identification and cloning of R-genes in Musa would contribute to germplasm improvement. To date, five main R-gene classes have been identified, based upon protein domains, with the most abundant coding for nucleotide-binding site-leucine-rich repeat (NBS-LRR) proteins. Primers designed from conserved protein motifs have enabled amplification of NBS homologues across diverse plant species. In Musa, our group has identified over 50 distinct NBS-LRR type resistance gene analogs (RGAs) in the resistant wild diploid M. acuminata Calcutta 4. The aim of this work was to characterize RGAs in M. acuminata cultivars contrasting in resistance to Black leaf Streak Disease. PCR amplification was conducted using DNA from M. acuminata cultivars Calcutta 4 (resistant) and Pisang Berlin (susceptible). Degenerate primers targeted sequences homologous to the NBS-LRR R-gene family. Following sequencing and processing of cloned PCR products, 63 out of a total of 136 high quality sequences showed homology to R-genes or RGAs. Phylogenetic analysis was conducted on deduced amino-acid sequences. Degenerate primers were also developed targeting an R-gene family of cytoplasmic serine-threonine (Ser/Thr) receptor-like kinases (RLKs) with extracellular LRRs, for application across cultivars. Studies are also planned for selection and full length sequencing of clones from M. acuminata and M. balbisiana BAC libraries containing novel RGAs characterized in this study, as an approach for complete R-gene sequence characterization, applicable both in transformation and breeding programs for banana genetic improvement. (author)