Cloning and characterization of a novel secretory root-expressed peroxidase gene from common bean (Phaseolus vulgaris L.) infected with Fusarium oxysporum f. sp. Phaseoli

Peroxidases play an important role in plant defense response to the attack of a host tissue by many pathogens. In this study, a new POX gene was isolated from common bean roots using cDNA-AFLP libraries and differential display of tissues with and without Fusarium oxysporum f. sp. phaseoli infection. Rapid amplification of cDNA ends was used to clone the full gene. The source of the differentially expressed fragment was the resistant genotype CAAS260205. The common bean POX gene showed a similar gene structure to a soybean POX gene consisting of four exons and three introns. Sequence analysis showed that PvPOX1 contained two conserved functional domains: a peroxidae active enzymatic site and a peroxidase heme-ligand site in close proximity. Phylogenetic study differentiated the new PvPOX1 gene from previous peroxidases in this multi-gene family. Real-time PCR analyses indicated that expression of the PvPOX1 gene was upregulated by Fusarium infection and that peroxidase activity, as measured by hydrogen peroxide (H2O2) accumulation, was enhanced during disease. The mRNA transcript level of PvPOX1 was also significantly increased following the stresses of wounding and polyethylene glycol, salicylic acid and abscisic acid treatments. The results of subcellular localization studies indicated that the localization of the PvPOX1 protein was in the apoplastic region of root cells. A co-dominant marker was developed for genetic mapping of the PvPOX1 gene based on an indel/insertion between the sequences of Andean and Mesoamerican parental genotypes (Hongyundou and Jingdou), which are mapping parents used in resistance breeding. Based on the new genetic marker, the gene was located between simple sequence repeat markers BMg2113 and BM181 on chromosome 3 of common bean and was not related to other peroxidases in terms of location. Overall, these results facilitate understanding of the molecular mechanism controlling resistance to Fusarium wilt pathogens and provide a diagnostic marker for selection of resistance to this disease based on peroxidase expression in common bean.