Cloning and Characterization of Resistance Gene Analogues (RGAs) from Piper Nigrum L. cv. Semongok Aman and Piper Colubrinum Link

Plant resistance gene analogues (RGAs) are known to share similarities in protein structural motifs. Cloning of RGAs in Piper spp. aims to offer potential in breeding for disease resistant varieties. A pair of degenerate primer designed from the conserved motifs of RGAs was used to amplify the Nucleotide-Triphosphate Binding Site (NBS) regions of P. nigrum cv. Semongok Aman and P. colubrinum. A total of ten partial RGAs designated as PcRGAt5, PcRGAt6, PcRGAt7, PcRGAt8, PcRGAt9, PnRGAt10, PnRGAt11, PnRGAt12, PnRGAt13 and PnRGAt14 have been sequenced. These RGAs range from 498 bp to 540 bp and encode proteins with 165 amino acids to 179 amino acids. The predicted RGA peptides contained the Kinase-1/P-loop (GMGGVGKT), Kinase-2 (VLDDVW) and hydrophobic Kinase-3/GLPL domains. These peptides show 42% to 47% similarity with the published NBS-type resistance proteins of Prunus spp., Malus baccata and Manihot esculenta. The clustering analysis has revealed that NBS-type resistance genes comprise a large gene family in Piper spp.