RNA 3′ uridylation occurs pervasively in eukaryotes, but is poorly characterized in viruses. In this study, we demonstrate that a broad array of RNA viruses, including mycoviruses, plant viruses and animal viruses, possess a novel population of RNA species bearing nontemplated oligo(U) or (U)-rich tails, suggesting widespread 3′ uridylation in eukaryotic viruses. Given the biological relevance of 3′ uridylation to eukaryotic RNA degradation, we propose a conserved but as-yet-unknown mechanism in virus-host interaction.
Background Alhagi sparsifolia is a typical desert phreatophyte and has evolved to withstand extreme dry, cold and hot weather. While A. sparsifolia represents an ideal model to study the molecular mechanism of plant adaption to abiotic stress, no research has been done in this aspect to date. Here we took advantage of Illumina platform to survey transcriptome in primary roots of A. sparsifolia under water stress conditions in aim to facilitate the exploration of its genetic basis for drought tolerance.Methodology and Principal FindingsWe sequenced four primary roots samples individually collected at 0, 6, 24 and 30h from the A. sparsifolia seedlings in the course of 24h of water stress following 6h of rehydration. The resulting 38,763,230, 67,511,150, 49,259,804 and 54,744,906 clean reads were pooled and assembled into 33,255 unigenes with an average length of 1,057 bp. All-unigenes were subjected to functional annotation by searching against the public databases. Based on the established transcriptome database, we further evaluated the gene expression profiles in the four different primary roots samples, and identified numbers of differently expressed genes (DEGs) reflecting the early response to water stress (6h vs. 0h), the late response to water stress (24h vs. 0h) and the response to post water stress rehydration (30h vs. 24h). Moreover, the DEGs specifically regulated at 6, 24 and 30h were captured in order to depict the dynamic changes of gene expression during water stress and subsequent rehydration. Functional categorization of the DEGs indicated the activation of oxidoreductase system, and particularly emphasized the significance of the ‘Glutathione metabolism pathway’ in response to water stress.ConclusionsThis is the first description of the genetic makeup of A. sparsifolia, thus providing a substantial contribution to the sequence resources for this species. The identified DEGs offer a deep insight into the molecular mechanism of A. sparsifolia in response to water stress, and merit further investigation.
Summary. -The occurrence of the primer-independent cDNA synthesis during RT-PCR analysis of human and animal RNA viruses has been well documented. Conversely, there is scant knowledge about this event in plant RNA viruses. Here we show that the primer-independent cDNA synthesis occurs in all eight different plant RNA viruses tested in this study, suggesting a common phenomenon for RT-PCR analysis of plant RNA viruses. Additional experiments indicate that the event is likely contributed to by RNA self-priming, and can be effectively reduced or eliminated through increasing temperature of the RT reaction.Keywords: RT-PCR; primer-independent cDNA synthesis; self-priming; plant RNA virus; RT temperature * Corresponding author. E-mail: liweimin01@caas.cn; phone: +86-10-82106117. C. Zhang. and H.N. Wu contributed equally to this work. Abbreviations: CGMMV = cucumber green mottle mosaic virus; CMV = cucumber mosaic virus; gRNA = genomic RNA; ORSV = odontoglossum ring-spot virus; PVX = potato virus X; TCV = turnip crinkle virus; TMV U1 = tobacco mosaic virus U1 strain; TNV = tobacco necrosis virus; TRV = tobacco rattle virus RT-PCR is one of the most common methods of molecular biology used to detect and quantify RNA expression levels in both cells and small quantities of tissues. Within this process, RT is the first and fundamental step, in which the RNA template is converted into its cDNA by reverse transcriptase in the presence of exogenous oligonucleotide referred to as primer, providing template DNA for subsequent PCR amplification. The primer-dependent mechanism for RT reaction has been generally accepted over the years. However, during RT-PCR analysis of human and animal RNA viruses, bacterial operons, as well as eukaryotic cellular RNAs, it has been observed that cDNA could be synthesized by reverse transcriptase without the addition of exogenous oligonucleotide (Gunji et al
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