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.
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:BackgroundLeucojum aestivum L. is an important medicinal plant which produces Amaryllidaceae alkaloids, especially galanthamine and lycorine. Research is currently exploring the possibility of producing these alkaloids using biotechnological methods, including in vitro cultures. The biosynthesis of alkaloids may be affected by the types and concentrations of carbohydrate sources used in the medium. In the present investigation we performed such studies on in vitro cultures of L. aestivum with a view to obtaining plant material of good quality, characterized, in particular, by a high content of valuable Amaryllidaceae alkaloids.MethodsWe examined the effects of various types of carbohydrate sources-sucrose, glucose, fructose and maltose-at different concentrations (30, 60 and 90 g/L)-on the quality of L. aestivum plants grown in the RITA® bioreactor. The plants' quality was assessed by their biomass increments, as well by as analysing photosynthetic pigments, endogenous sugar, phenolics and Amaryllidaceae alkaloid content. We also investigated the effect of sugars on the activity of the antioxidant enzymes catalase (CAT), peroxidase (POD) and superoxide dismutase (SOD).ResultsThe highest biomass increments were observed in plants cultivated in the medium containing 90 g/L sucrose. The highest CAT activity was noted in cultures growing in the medium supplemented with 90 g/L maltose, while the highest POD activity was observed in the presence of 90 g/L fructose and 60 g/L maltose. No differences in SOD activity were observed. Moreover, the sugars did not affect the contents of chlorophyll a and carotenoids, whereas the highest amount of chlorophyll b was recorded in plants growing in the medium with 60 g/L maltose. No statistically significant differences were observed in the contents of endogenous sugars and phenolics in any in vitro conditions. However, the addition of sugar had a decisive effect on the biosynthesis of the Amaryllidaceae alkaloids. The highest distribution of alkaloids occurred in plants cultured in the medium containing 60 g/L sucrose. Six Amaryllidaceae alkaloids were detected in the plant tissue. The addition of 30 g/L fructose in the medium resulted in the accumulation of five alkaloids, including ismine, which was not identified in other analysed tissues. The highest concentration of galanthamine was observed in plants cultured in the presence of 30 g/L fructose and 60 g/L sucrose (39.2 and 37.5 µg/g of dry weight (DW), respectively). The plants grown in the medium containing 60 g/L sucrose exhibited the highest lycorine content (1048 µg/g of DW).ConclusionsThe type and concentration of sugar used in the medium have an essential influence on the biosynthesis of Amaryllidaceae alkaloids in L. aestivum plants cultured in a RITA® bioreactor. The results point to an interesting approach for commercial production of galanthamine and lycorine.