Transcriptome analysis reveals key defense related differentially expressed genes in response to stripe rust on wheat

A near isogenic line FLW29 containing yellow rust resistance gene Yr16 introgressed from wild wheat species (Cappelle-Desprez) was crossed with wheat cultivar PBW343. The recipient parent FLW29 (resistant), and cv. PBW343 (susceptible) were used for studying the differential gene expression in response to Puccinia striiformis f. sp. tritici (Pst). Two wheat genotypes were inoculated with Pst pathotype 46S119 such that for each cultivar a total of 10 pots (5 mock-inoculated and 5 inoculated) with three biological replicates were used for the experiment. Fresh uredinospores of 46S119 were harvested from infected wheat plant and resuspended in sterile distilled water evenly mixed with soltrol (20mg/100ml) that was used to inoculate seedlings growth. Seedlings at two-leaf stage/flag leaves (approx 20 days after sowing), were inoculated with the spore suspension using an automizer and then kept in dark at 10 degree C for 16-h light/8-hdark to maintain the relative high humidity. Later, leaf samples were collected at 3 different time periods i,e. at 12, 48 and 72 hr post-inoculation (HPI). The collected samples were immediately placed in RNA later and stored at -20 degree C until use. The RNA from leaf samples were extracted using Qiagen RNeasy Mini kit (QiagenInc, USA). Samples were IIumina sequenced on a single HiSeq 4000 machines yielding at least 895M 150bp paired-end reads per sample. Each sample represents one experimental condition (pathogen/mock; time points; genotype).