Identification of candidate genes associated with CBB resistance in common bean HR45 (Phaseolus vulgaris L.) using cDNA-AFLP

Abstract

Common bacterial blight (CBB), incited by Xanthomonas axonopodis pv. phaseoli (Xap), is a serious seed-borne disease of common bean (Phaseolus vulgaris L.) in both temperate and tropical production zones. The line HR45 is highly resistant to Xap infection on leaves and pods in both field and greenhouse. To understand the molecular mechanisms underlying CBB resistance in HR45, cDNA-amplified fragment length polymorphism (AFLP) technique was used to identify the genes that are differentially expressed in the leaves of HR45 at different time-periods after inoculation. Selective amplifications with 34 primer combinations allowed the visualization of 2,448 transcript-derived fragments (TDFs) in infected leaves, and 259 (10.6%) of them were differentially expressed TDFs (DE-TDFs). Seventy-seven of the DE-TDFs were cloned and sequenced. Thirty-nine of the 77 (50.6%) DE-TDFs representing bean transcripts were not previously reported in any EST database. The expression patterns of 10 representative DE-TDFs were further confirmed by real-time RT-PCR. BLAST analysis suggested that 40% (31 of 77) of the DE-TDFs were homologous to the genes related to metabolism, photosynthesis, and cellular transport, whereas 28% (22 of 77) of the DE-TDFs showed homology to the genes involved in defence response, response to stimulus, enzyme regulation, and transcription regulation. Thus, the 22 pathogenesis-related DE-TDFs were selected as potential functional candidate genes (FCGs) in association with CBB resistance. Meanwhile, six of the DE-TDFs (1FCG and five other DE-TDFs) were in silico mapped to the distal region of the bean linkage group B6 (the genomic location containing the major CBB resistance QTL in HR45) and, therefore, were considered as positional candidate genes (PCGs). This study represents a first step towards the discovery of bean genes expressed upon Xap infection. This information will be useful for elucidating the molecular basis of the resistance response process and identifying the genes that underlie the CBB-resistance.