Identification of differentially expressed genes in sugarcane during pathogenesis of Colletotrichum falcatum by suppression subtractive hybridization (SSH)

Abstract

A suppression subtractive hybridization (SSH) technique was used to identify differentially expressed genes in sugarcane in response to Colletotrichum falcatum, the fungal pathogen causing red rot in sugarcane. Messenger RNA (mRNA) samples collected from red rot resistant cv. Co 93009 at 6, 12, 24 and 48h after pathogen challenge were pooled together and used as tester population. mRNA samples collected from red rot susceptible cv. CoC 671 at 6, 12, 24 and 48h after pathogen challenge and control of samples of cv. Co 93009 and cv. CoC 671 were pooled together and used as the driver population. A forward subtraction enriched for differentially expressed ESTs in the resistant cultivar was carried out. At the end of subtraction, cloning and sequencing, 136 EST sequences were assembled into 10 clusters/contigs. Based on TIGR homology search, the clusters were found to be involved in reactive oxygen species (ROS) signaling, defense and the secretory pathway of plant innate immunity associated with hypersensitive response (HR) - mediated programmed cell death (PCD). Temporal gene expression pattern of the EST clusters by quantitative real time PCR (qRT-PCR) revealed that, resistant cultivar had a higher level of gene expression than the susceptible cultivar in all the clusters till 48h after pathogen inoculation. In conclusion, this study identified for the first time a set of differentially expressed EST clusters in red rot resistant sugarcane cultivar in response to C. falcatum infection