GSBS Student Publications

Student Author(s)

Erin E. Heyer

GSBS Program

Biochemistry & Molecular Pharmacology

UMMS Affiliation

Department of Biochemistry and Molecular Pharmacology; RNA Therapeutics Institute

Date

1-2015

Document Type

Article

Medical Subject Headings

DNA, Circular; DNA, Single-Stranded; Electrophoresis, Polyacrylamide Gel; *Gene Library; High-Throughput Nucleotide Sequencing; MicroRNAs; Reverse Transcriptase Polymerase Chain Reaction; Sequence Analysis, RNA

Disciplines

Biochemistry | Bioinformatics | Computational Biology | Genetics and Genomics | Nucleic Acids, Nucleotides, and Nucleosides

Abstract

Deep sequencing of strand-specific cDNA libraries is now a ubiquitous tool for identifying and quantifying RNAs in diverse sample types. The accuracy of conclusions drawn from these analyses depends on precise and quantitative conversion of the RNA sample into a DNA library suitable for sequencing. Here, we describe an optimized method of preparing strand-specific RNA deep sequencing libraries from small RNAs and variably sized RNA fragments obtained from ribonucleoprotein particle footprinting experiments or fragmentation of long RNAs. Our approach works across a wide range of input amounts (400 pg to 200 ng), is easy to follow and produces a library in 2-3 days at relatively low reagent cost, all while giving the user complete control over every step. Because all enzymatic reactions were optimized and driven to apparent completion, sequence diversity and species abundance in the input sample are well preserved.

Rights and Permissions

Citation: Nucleic Acids Res. 2015 Jan;43(1):e2. doi: 10.1093/nar/gku1235. Epub 2014 Dec 12. Link to article on publisher's site

DOI of Published Version

10.1093/nar/gku1235

Comments

© The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited.

Related Resources

Link to Article in PubMed

Journal Title

Nucleic acids research

PubMed ID

25505164

Creative Commons License

Creative Commons Attribution-Noncommercial 4.0 License
This work is licensed under a Creative Commons Attribution-Noncommercial 4.0 License

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