A comparative study of ChIP-seq sequencing library preparation methods

BMC Genomics. 2016 Oct 21;17(1):816. doi: 10.1186/s12864-016-3135-y.


Background: ChIP-seq is the primary technique used to investigate genome-wide protein-DNA interactions. As part of this procedure, immunoprecipitated DNA must undergo "library preparation" to enable subsequent high-throughput sequencing. To facilitate the analysis of biopsy samples and rare cell populations, there has been a recent proliferation of methods allowing sequencing library preparation from low-input DNA amounts. However, little information exists on the relative merits, performance, comparability and biases inherent to these procedures. Notably, recently developed single-cell ChIP procedures employing microfluidics must also employ library preparation reagents to allow downstream sequencing.

Results: In this study, seven methods designed for low-input DNA/ChIP-seq sample preparation (Accel-NGS® 2S, Bowman-method, HTML-PCR, SeqPlex™, DNA SMART™, TELP and ThruPLEX®) were performed on five replicates of 1 ng and 0.1 ng input H3K4me3 ChIP material, and compared to a "gold standard" reference PCR-free dataset. The performance of each method was examined for the prevalence of unmappable reads, amplification-derived duplicate reads, reproducibility, and for the sensitivity and specificity of peak calling.

Conclusions: We identified consistent high performance in a subset of the tested reagents, which should aid researchers in choosing the most appropriate reagents for their studies. Furthermore, we expect this work to drive future advances by identifying and encouraging use of the most promising methods and reagents. The results may also aid judgements on how comparable are existing datasets that have been prepared with different sample library preparation reagents.

Keywords: Chromatin immunoprecipitation; HTS; Low-input; Micro-ChIP; NGS.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Chromatin Immunoprecipitation* / methods
  • Chromosome Mapping
  • Gene Library*
  • Genome
  • Genomics / methods
  • High-Throughput Nucleotide Sequencing* / methods
  • Humans
  • Reproducibility of Results
  • Sequence Analysis, DNA