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Deciphering the RNA landscape by RNAome sequencing [Seq]

ABSTRACT: Current RNA expression profiling methods rely on enrichment steps for specific RNA classes, thereby not detecting all RNA species in an unperturbed manner. We report strand-specific RNAome sequencing that determines expression of small and large RNAs from ribosomal RNA-depleted total RNA in a single sequence run. Since current analysis pipelines cannot reliably analyze small and large RNAs simultaneously, we developed TRAP, Total Rna Analysis Pipeline, a robust interface that is also compatible with existing RNA sequencing protocols. RNAome sequencing quantitatively preserved all RNA classes, allowing cross-class comparisons that facilitates the identification of relationships between different RNA classes. We demonstrate the strength of RNAome sequencing in mouse embryonic stem cells treated with cisplatin. MicroRNA and mRNA expression in RNAome sequencing significantly correlated between replicates and was in concordance with both existing RNA sequencing methods and gene expression arrays generated from the same samples. Moreover, RNAome sequencing also detected additional RNA classes such as enhancer RNAs, anti-sense RNAs, novel RNA species and numerous differentially expressed RNAs undetectable by other methods. At the level of complete RNA classes, RNAome sequencing also identified a specific global repression of the microRNA and microRNA isoform classes after cisplatin treatment whereas all other classes such as mRNAs were unchanged. These characteristics of RNAome sequencing will significantly improve expression analysis as well as studies on RNA biology not covered by existing methods.

ORGANISM(S): Mus musculus

PROVIDER: GSE48083 | GEO | 2015/04/01

SECONDARY ACCESSION(S): PRJNA208891

REPOSITORIES: GEO

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Project description:Melon (Cucumis melo L.) is a commercially important fruit crop that is cultivated worldwide. The melon research community has recently benefited from the determination of a complete draft genome sequence and the development of associated genomic tools, which have allowed us to focus on small RNAs (sRNAs). These are short, non-coding RNAs 21â24 nucleotides in length with diverse physiological roles. In plants, they regulate gene expression and heterochromatin assembly, and control protection against virus infection. Much remains to be learned about the role of sRNAs in melon. We constructed 10 sRNA libraries from two stages of developing ovaries, fruits and photosynthetic cotyledons infected with viruses, and carried out high-throughput pyrosequencing. We catalogued and analyzed the melon sRNAs, resulting in the identification of 26 known miRNA families (many conserved with other species), the prediction of 84 melon-specific miRNA candidates, the identification of trans-acting siRNAs, and the identification of chloroplast, mitochondrion and transposon-derived sRNAs. In silico analysis revealed more than 400 potential targets for the conserved and novel miRNAs. This analysis provides insight into the composition and function of the melon small RNAome, and paves the way towards an understanding of sRNA-mediated processes that regulate melon fruit development and melonâvirus interactions. 11 small RNA libraries from several tissues of melon are included en the raw data. 2 samples from ovary, 2 samples from fruit, 1 sample from healthy cotyledons (Cultivar Tendral), 1 samples from healthy cotyledons (genotype TGR-1551), 1 sample from cotyledons (cultivar Tendral) infected with Watermelon mosaic virus (WMV), 1 sample from cotyledons (cultivar TGR-1551) infected with WMV, 1 sample from cotyledons (cultivar Tendral) infected with Melon necrotic spot virus (MNSV, Malfa5 isolate), 1 sample from cotyledons (cultivar Tendral) infected with MNSV (chimeric virus with Malfa5-264 isolates), 1 library from synthetic RNA oligos. Raw reads were obtained from two independent 454 runs, ~22,000 reads each one, to a total of 447,180 reads

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Deciphering the RNA landscape by RNAome sequencing [Seq]

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