NanoPARE: parallel analysis of RNA 5' ends from low-input RNA.

Genome Res 2018 12 24;28(12):1931-1942. Epub 2018 Oct 24.

Gregor Mendel Institute (GMI), Austrian Academy of Sciences, Vienna Biocenter (VBC), 1030 Vienna, Austria.

Diverse RNA 5' ends are generated through both transcriptional and post-transcriptional processes. These important modes of gene regulation often vary across cell types and can contribute to the diversification of transcriptomes and thus cellular differentiation. Therefore, the identification of primary and processed 5' ends of RNAs is important for their functional characterization. Methods have been developed to profile either RNA 5' ends from primary transcripts or the products of RNA degradation genome-wide. However, these approaches either require high amounts of starting RNA or are performed in the absence of paired gene-body mRNA-seq data. This limits current efforts in RNA 5' end annotation to whole tissues and can prevent accurate RNA 5' end classification due to biases in the data sets. To enable the accurate identification and precise classification of RNA 5' ends from standard and low-input RNA, we developed a next-generation sequencing-based method called nanoPARE and associated software. By integrating RNA 5' end information from nanoPARE with gene-body mRNA-seq data from the same RNA sample, our method enables the identification of transcription start sites at single-nucleotide resolution from single-cell levels of total RNA, as well as small RNA-mediated cleavage events from at least 10,000-fold less total RNA compared to conventional approaches. NanoPARE can therefore be used to accurately profile transcription start sites, noncapped RNA 5' ends, and small RNA targeting events from individual tissue types. As a proof-of-principle, we utilized nanoPARE to improve RNA 5' end annotations and quantify microRNA-mediated cleavage events across five different flower tissues.

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http://dx.doi.org/10.1101/gr.239202.118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6280765PMC
December 2018
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References

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Genetic interactions among floral homeotic genes of Arabidopsis
Development 1991

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