Project description:The late blight pathogen, Phytophthora infestans has a broad host range within the Solanaceae family, including yellow potato (Solanum phureja). The disease caused by P. infestans in S. phureja is poorly understood and is a major concern in Colombia. Expressed Sequence Tag (EST) libraries obtained from a normalized library constructed from healthy plant tissue revealed high levels of sequence similarity between S. phureja and S. tuberosum. Then, utilizing Serial Analysis of Gene Expression and high-throughput sequencing (SAGE-Solexa), we characterized yellow potato gene expression during infection by P. infestans. Four-week-old yellow potato plants were inoculated with P. infestans and were collected at 12 and 72 hours post inoculation for RNA extraction. We detected differentially expressed genes by comparing inoculated to non-inoculated and resistant to susceptible plants. The discovery and characterization of the proteins mediating this host–pathogen interaction enable the understanding of the pathosystem and is the key for developing resistant plants. Keywords: SAGE-Solexa, inoculation response, transcript profiling, Solanum phureja, Phytophthora infestans Four-week-old yellow potato (Solanum phureja) plants were inoculated with Phytophthora infestans and were collected and flash frozen in liquid nitrogen at 12 and 72 hours post inoculation, as well as mock inoculated, for RNA extraction. 2 yellow potato cultivars (resistant and susceptible) were used for each experiment. Mock inoculated plants were collected in each replicate. RNA obtained from each of the three biological replicates was pooled to obtain a single RNA sample for each timepoint X cultivar combination. A total of 6 different SAGE libraries were thus obtained. For all libraries, Illumina sequencing was performed at Canada´s Michael Smith Genome Sciences Centre.
Project description:Identification of novel potato candidate genes for quantitative resistance to Phytophthora infestans by SuperSAGE transcriptome analysis
Project description:The late blight pathogen, Phytophthora infestans has a broad host range within the Solanaceae family, including yellow potato (Solanum phureja). The disease caused by P. infestans in S. phureja is poorly understood and is a major concern in Colombia. Expressed Sequence Tag (EST) libraries obtained from a normalized library constructed from healthy plant tissue revealed high levels of sequence similarity between S. phureja and S. tuberosum. Then, utilizing Serial Analysis of Gene Expression and high-throughput sequencing (SAGE-Solexa), we characterized yellow potato gene expression during infection by P. infestans. Four-week-old yellow potato plants were inoculated with P. infestans and were collected at 12 and 72 hours post inoculation for RNA extraction. We detected differentially expressed genes by comparing inoculated to non-inoculated and resistant to susceptible plants. The discovery and characterization of the proteins mediating this host–pathogen interaction enable the understanding of the pathosystem and is the key for developing resistant plants. Keywords: SAGE-Solexa, inoculation response, transcript profiling, Solanum phureja, Phytophthora infestans
Project description:Phytophthora spp. encode large sets of effector proteins and distinct populations of small RNAs (sRNAs). Reports suggest that pathogen-derived sRNAs can modulate the expression of plant defense genes. The experiments reported here were designed to shed light on impact of sRNAs in the potato-P. infestans interaction. We used the Argonaute or Ago1 from P. infestans tagged with GFP transformed into the 88069 strain to infect potato cv. Bintje plants. Collected leaf materials were used in co-immunoprecipitation experiments together with P. infestans harboring GFP (control GFP) and P. infestans mycelia grown on media (control mycelia). These three materials were sequenced at a Ion Proton platform. The reads length of 8-38 nt were adaptor-trimmed and mapped to the P. infestans genome and the Solanom tuberosum genome v4.04. Both P. infestans-associated and potato derived sRNAs were identified.
Project description:Nitrogen (N) fertilization is an important abiotic factor for the growth of potato (S. tuberosum) because of its potential effects on yield. Because excess N in the soil runs off into water systems and negatively impacts the environment, studies on N use by the plant are key to decrease N-fertilizer use. Three commercial potato cultivars (Shepody, Russet-Burbank and Atlantic) were grown under two different rates of applied N-fertilizer (0 kg N ha-1 and 180 kg N ha-1) to obtain more information on the underlying gene regulation mechanisms associated with N. Plants with no added N had significantly lower concentrations of petiole nitrates, chlorophyll level indices, biomass and yield per hectare. Total mRNA samples were taken at two different time-points during the growth season and used for sequencing. The results for each cultivar and time-point were analysed separately to find differentially expressed genes. In total, thirty genes were found to be over-expressed and nine genes were found to be under-expressed in plants from all potato cultivars when they were grown with added N-fertilizer. The 1000 bp upstream flanking regions of the differentially expressed genes were analysed to find overrepresented motifs using three motif discovery algorithms (Seeder, Weeder and MEME). Nine different motifs were found, indicating potential gene regulatory mechanisms for potato under N-deficiency.
Project description:Potato (Solanum tuberosum) late blight disease caused by Phytophthora. infestans is one of the most critical crop diseases in the world. We investigated the proteome of Solanum tubersoum (cv. Sarpo Mira) by quantitative proteomics after foliar application of encysted zoospore suspension from P. infestans isolate. Also, we examined changes in protein abundance of cv. Sarpo Mira in response to P. infestans infection at three key timepoints 0 hpi (L), 48 hpi (M) and 120 hpi (H), divided into early and late disease stages and analyzed the protein-protein interaction during the disease stages. In response to P. infestans oomycete infection, the iTRAQ comparative proteome survey revealed a total of 1229 differentially expressed proteins (DEPs) in the pairwise comparison of disease stages, which included DEPs specific to early disease stage, DEPs specific to late disease stage and commonly shared DEPs. Over 80% of the change in protein abundance were up-regulated in the early stages of infection while about 61% of the DEPs in the later disease stage were down-regulated. The analysis of expression patterns, functional category, and enrichment tests highlighted significant coordination and enrichment of cell wall-associated defense response proteins in the early stages and cellular protein modification process as well as membrane protein complex formation in the late stages. Furthermore, strong protein interactions were observed in the late disease stage that might relate to disease pathogenesis.