Monday, 10 February 2014

PubMed Highlights: NGS library prepartion

Starting with a robust sequencing library is the first and crucial step to obtain unbiased, high-quality results from your next generation sequencher!
Here are a couple of interesting paper reviewing problems and solutions related to the NGS library preparation. They also give a sinthetic overview on the present library prepartion methods and how they fit to different downstream applications.
Take a look!

Library preparation methods for next-generation sequencing: Tone down the bias.
Exp Cell Res. 2014 Jan 15

Next-generation sequencing (NGS) has caused a revolution in biology. NGS requires the preparation of libraries in which (fragments of) DNA or RNA molecules are fused with adapters followed by PCR amplification and sequencing. It is evident that robust library preparation methods that produce a representative, non-biased source of nucleic acid material from the genome under investigation are of crucial importance. Nevertheless, it has become clear that NGS libraries for all types of applications contain biases that compromise the quality of NGS datasets and can lead to their erroneous interpretation. A detailed knowledge of the nature of these biases will be essential for a careful interpretation of NGS data on the one hand and will help to find ways to improve library quality or to develop bioinformatics tools to compensate for the bias on the other hand. In this review we discuss the literature on bias in the most common NGS library preparation protocols, both for DNA sequencing (DNA-seq) as well as for RNA sequencing (RNA-seq). Strikingly, almost all steps of the various protocols have been reported to introduce bias, especially in the case of RNA-seq, which is technically more challenging than DNA-seq. For each type of bias we discuss methods for improvement with a view to providing some useful advice to the researcher who wishes to convert any kind of raw nucleic acid into an NGS library.

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.

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