Performance of microarray and liquid based capture methods for target enrichment for massively parallel sequencing and SNP discovery

Abstract

Targeted sequencing is a cost-efficient way to obtain answers to biological questions in many projects, but the choice of the enrichment method to use can be difficult. In this study we compared two hybridization methods for target enrichment for massively parallel sequencing and single nucleotide polymorphism (SNP) discovery, namely Nimblegen sequence capture arrays and the SureSelect liquid-based hybrid capture system. We prepared sequencing libraries from three HapMap samples using both methods, sequenced the libraries on the Illumina Genome Analyzer, mapped the sequencing reads back to the genome, and called variants in the sequences. 74-75% of the sequence reads originated from the targeted region in the SureSelect libraries and 41-67% in the Nimblegen libraries. We could sequence up to 99.9% and 99.5% of the regions targeted by capture probes from the SureSelect libraries and from the Nimblegen libraries, respectively. The Nimblegen probes covered 0.6 Mb more of the original 3.1 Mb target region than the SureSelect probes. In each sample, we called more SNPs and detected more novel SNPs from the libraries that were prepared using the Nimblegen method. Thus the Nimblegen method gave better results when judged by the number of SNPs called, but this came at the cost of more over-sampling.

Figure 1. Coverage plots of the sequencing libraries prepared with SureSelect (left) and Nimblegen (right) methods.

Cumulative base frequency is plotted against coverage. In the SureSelect libraries, 94–97% of the bases in the regions targeted by the probes are covered by at least ten reads. In the Nimblegen libraries the corresponding number is 83–97%.

Figure 2. Coverage of a single gene.

Per base coverage plotted along ARL11 in the sample NA11993 in sequencing libraries prepared with Nimblegen (top) and SureSelect (bottom) methods. The bars at the top of the graph mark the regions that are targeted by probes in each design (red) and the repeat elements as determined by RepeatMasker (blue).

Figure 3. Normalized coverage of the sequencing libraries prepared with SureSelect (left) and Nimblegen (right) methods.

To normalize the coverage, the absolute per base coverage was divided by the mean coverage. In the SureSelect libraries, 67–69% of the regions targeted by probes are covered by at least half of the average coverage. In the Nimblegen libraries 61–72% of the regions targeted by probes are covered by at least half of the average coverage.