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Phospho-RNA-seq: a modified small RNA-seq method that reveals circulating mRNA and lncRNA fragments as potential biomarkers in human plasma

AuthorsGiráldez, María D.; Spengler, Ryan M.; Etheridge, Alton; Goicochea, Annika J.; Tuck, Missy; Won Choi, Sung; Galas, David J.; Tewari, Muneesh
Issue Date3-Jun-2019
PublisherEMBO Press
CitationEMBO Journal 38(11): e101695 (2019)
AbstractExtracellular RNA s (exRNA s) in biofluids have attracted great interest as potential biomarkers. Although extracellular microRNA s in blood plasma are extensively characterized, extracellular messenger RNA (mRNA ) and long non‐coding RNA (lncRNA ) studies are limited. We report that plasma contains fragmented mRNA s and lncRNA s that are missed by standard small RNA ‐seq protocols due to lack of 5′ phosphate or presence of 3′ phosphate. These fragments were revealed using a modified protocol (“phospho‐RNA ‐seq”) incorporating RNA treatment with T4‐polynucleotide kinase, which we compared with standard small RNA ‐seq for sequencing synthetic RNA s with varied 5′ and 3′ ends, as well as human plasma exRNA . Analyzing phospho‐RNA ‐seq data using a custom, high‐stringency bioinformatic pipeline, we identified mRNA /lncRNA transcriptome fingerprints in plasma, including tissue‐specific gene sets. In a longitudinal study of hematopoietic stem cell transplant patients, bone marrow‐ and liver‐enriched exRNA genes were tracked with bone marrow recovery and liver injury, respectively, providing proof‐of‐concept validation as a biomarker approach. By enabling access to an unexplored realm of mRNA and lncRNA fragments, phospho‐RNA ‐seq opens up new possibilities for plasma transcriptomic biomarker development.
Publisher version (URL)http://dx.doi.org/10.15252/embj.2019101695
Identifierse-issn: 1460-2075
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