Exome sequencing from nanogram amounts of starting DNA: comparing three approaches

Vera N Rykalina; Alexey A Shadrin; Vyacheslav S Amstislavskiy; Evgeny I Rogaev; Hans Lehrach; Tatiana A Borodina

doi:10.1371/journal.pone.0101154

Exome sequencing from nanogram amounts of starting DNA: comparing three approaches

PLoS One. 2014 Jul 3;9(7):e101154. doi: 10.1371/journal.pone.0101154. eCollection 2014.

Authors

Vera N Rykalina¹, Alexey A Shadrin², Vyacheslav S Amstislavskiy³, Evgeny I Rogaev⁴, Hans Lehrach³, Tatiana A Borodina⁵

Affiliations

¹ Max-Planck Institute for Molecular Genetics, Berlin, Germany; AlacrisTheranostics GmbH, Berlin, Germany; Freie Universität Berlin, Berlin, Germany.
² Max-Planck Institute for Molecular Genetics, Berlin, Germany; Freie Universität Berlin, Berlin, Germany.
³ Max-Planck Institute for Molecular Genetics, Berlin, Germany.
⁴ Vavilov Institute of General Genetics, Moscow, Russia.
⁵ Max-Planck Institute for Molecular Genetics, Berlin, Germany; AlacrisTheranostics GmbH, Berlin, Germany.

Abstract

Hybridization-based target enrichment protocols require relatively large starting amounts of genomic DNA, which is not always available. Here, we tested three approaches to pre-capture library preparation starting from 10 ng of genomic DNA: (i and ii) whole-genome amplification of DNA samples with REPLI-g (Qiagen) and GenomePlex (Sigma) kits followed by standard library preparation, and (iii) library construction with a low input oriented ThruPLEX kit (Rubicon Genomics). Exome capture with Agilent SureSelectXT2 Human AllExon v4+UTRs capture probes, and HiSeq2000 sequencing were performed for test libraries along with the control library prepared from 1 µg of starting DNA. Tested protocols were characterized in terms of mapping efficiency, enrichment ratio, coverage of the target region, and reliability of SNP genotyping. REPLI-g- and ThruPLEX-FD-based protocols seem to be adequate solutions for exome sequencing of low input samples.

Publication types

Comparative Study
Research Support, Non-U.S. Gov't

MeSH terms

DNA / blood
DNA / genetics*
Exome*
Gene Library
Genomics / methods
Genotyping Techniques / methods
Humans
Sample Size
Sequence Analysis, DNA / methods*

Substances

DNA

Grants and funding

This work was supported by the 7th framework programme of the European Union (ADAMS) [242257, FP7-HEALTH-2009]. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.