@ARTICLE{10.3389/fpls.2015.00366, AUTHOR={Townsley, Brad T. and Covington, Michael F. and Ichihashi, Yasunori and Zumstein, Kristina and Sinha, Neelima R.}, TITLE={BrAD-seq: Breath Adapter Directional sequencing: a streamlined, ultra-simple and fast library preparation protocol for strand specific mRNA library construction}, JOURNAL={Frontiers in Plant Science}, VOLUME={6}, YEAR={2015}, URL={https://www.frontiersin.org/articles/10.3389/fpls.2015.00366}, DOI={10.3389/fpls.2015.00366}, ISSN={1664-462X}, ABSTRACT={Next Generation Sequencing (NGS) is driving rapid advancement in biological understanding and RNA-sequencing (RNA-seq) has become an indispensable tool for biology and medicine. There is a growing need for access to these technologies although preparation of NGS libraries remains a bottleneck to wider adoption. Here we report a novel method for the production of strand specific RNA-seq libraries utilizing the terminal breathing of double-stranded cDNA to capture and incorporate a sequencing adapter. Breath Adapter Directional sequencing (BrAD-seq) reduces sample handling and requires far fewer enzymatic steps than most available methods to produce high quality strand-specific RNA-seq libraries. The method we present is optimized for 3-prime Digital Gene Expression (DGE) libraries and can easily extend to full transcript coverage shotgun (SHO) type strand-specific libraries and is modularized to accommodate a diversity of RNA and DNA input materials. BrAD-seq offers a highly streamlined and inexpensive option for RNA-seq libraries.} }