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Biotechniques. 2014 Feb 1;56(2):61-4, 66, 68, passim. doi: 10.2144/000114133. eCollection 2014.

Library construction for next-generation sequencing: overviews and challenges.

Author information

1
NGS and Microarray Core Facility, The Scripps Research Institute, La Jolla, CA.
2
Department of Molecular and Experimental Medicine, The Scripps Research Institute, La Jolla, CA.
3
Laboratory of Pharmaceutical Biotechnology, Faculty of Pharmaceutical Sciences, Ghent University, Ghent, Belgium.

Abstract

High-throughput sequencing, also known as next-generation sequencing (NGS), has revolutionized genomic research. In recent years, NGS technology has steadily improved, with costs dropping and the number and range of sequencing applications increasing exponentially. Here, we examine the critical role of sequencing library quality and consider important challenges when preparing NGS libraries from DNA and RNA sources. Factors such as the quantity and physical characteristics of the RNA or DNA source material as well as the desired application (i.e., genome sequencing, targeted sequencing, RNA-seq, ChIP-seq, RIP-seq, and methylation) are addressed in the context of preparing high quality sequencing libraries. In addition, the current methods for preparing NGS libraries from single cells are also discussed.

KEYWORDS:

ChIP-seq; DNA; DNA-seq; RIP-seq; RNA; RNA-seq; deep sequencing; library preparation; next-generation sequencing

PMID:
24502796
PMCID:
PMC4351865
DOI:
10.2144/000114133
[Indexed for MEDLINE]
Free PMC Article

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