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. 2019 Feb 19;9:2262. doi: 10.1038/s41598-018-38458-7

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© The Author(s) 2019

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Modifications to high-throughput sequencing method improves capture of miRNAs. (A) Schematic of protocol to prepare miRNA libraries for sequencing. Modifications from original protocol noted in bold. (B) Percentage of different classes of RNAs captured from a plasma sample using the original conditions (0.85 μM 3′ adapter, 3.3 μM unmodified 5′ adapter). Ligations were performed in triplicate from the same RNA. Each replicate is shown as an individual bar. Note the low percentage of reads mapping to miRNAs (red). (C) Percentage of mature miRNAs captured using the optimized conditions (0.05 μΜ 3′ adapter, 0.33 μΜ amino-modified 5′ adapter) were compared to original conditions in three independent biological samples. The ligation reactions were performed in triplicate for each sample and protocol. Each replicate is shown as a black dot. Red dot represents average. (D) The average percentage of reads mapping to different classes of RNA for each sample and condition shown in C.