Project description:Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to compare NGS-derived Triticum aestivum transcriptome (RNA-seq) profiling methods and to evaluate genotypes associated with resistance against the Wheat dwarf virus. Methods: Triticum aestivum mRNA profiles of genotypes associated with resistance against the Wheat dwarf virus were generated by deep sequencing, in four replicates, using Illumina. The sequence reads that passed quality filters were analyzed at the transcript isoform level with two methods: Burrows–Wheeler Aligner (BWA) followed by ANOVA (ANOVA) and TopHat followed by Cufflinks. qRT–PCR validation was performed using TaqMan and SYBR Green assays. Conclusions: Our study represents the first detailed analysis of Triticum aestivum transcriptomes, with biologic replicates, generated by RNA-seq technology. The optimized data analysis workflows reported here should provide a framework for comparative investigations of expression profiles. Our results show that NGS offers a comprehensive and more accurate quantitative and qualitative evaluation of mRNA and miRNA content within a cell or tissue. We conclude that RNA-seq based transcriptome characterization would expedite genetic network analyses and permit the dissection of complex biologic functions.
Project description:We identified the long non-coding RNAs (lncRNAs) in Triticum aestivum infected with Fusarium graminearum by high-throughput RNA sequencing. More than 393 million clean reads were obtained from Illumina Hiseq 4000 system and 126,391 transcripts was identified as high-confidence lncRNAs in T. aestivum against F. graminearum by an integrated approach. Already well over 4,130 of the total 4,276 differentially expressed lncRNAs were specifically expressed at 12 h post-inoculation (hpi), but only 89 of these were specifically expressed at 24 hpi, indicating that the initial stage was the crucial stage for lncRNA-mediated gene regulation of wheat defense against F. graminearum. Target analysis showed the lncRNAs participated in various biological stress processes.
