Fosmid-based whole genome haplotyping of a HapMap trio child: evaluation of Single Individual Haplotyping techniques

Abstract

Determining the underlying haplotypes of individual human genomes is an essential, but currently difficult, step toward a complete understanding of genome function. Fosmid pool-based next-generation sequencing allows genome-wide generation of 40-kb haploid DNA segments, which can be phased into contiguous molecular haplotypes computationally by Single Individual Haplotyping (SIH). Many SIH algorithms have been proposed, but the accuracy of such methods has been difficult to assess due to the lack of real benchmark data. To address this problem, we generated whole genome fosmid sequence data from a HapMap trio child, NA12878, for which reliable haplotypes have already been produced. We assembled haplotypes using eight algorithms for SIH and carried out direct comparisons of their accuracy, completeness and efficiency. Our comparisons indicate that fosmid-based haplotyping can deliver highly accurate results even at low coverage and that our SIH algorithm, ReFHap, is able to efficiently produce high-quality haplotypes. We expanded the haplotypes for NA12878 by combining the current haplotypes with our fosmid-based haplotypes, producing near-to-complete new gold-standard haplotypes containing almost 98% of heterozygous SNPs. This improvement includes notable fractions of disease-related and GWA SNPs. Integrated with other molecular biological data sets, this phase information will advance the emerging field of diploid genomics.

HapCol: accurate and memory-efficient haplotype assembly from long reads

“…In fact, the advent of next-generation sequencing (NGS) technologies provided a cost-effective way of assembling the genome of diploid organisms. However, to assemble accurate haplotypes, it is necessary to have reads that are long enough to span several different heterozygous positions (Duitama et al, 2012). This kind of data is becoming increasingly available with the advent of 'future-generation' sequencing technologies such as single molecule real-time technologies like PacBio RS II (http:// www.pacificbiosciences.com/products/) and Oxford Nanopore flow cell technologies like MinION (https://www.nanoporetech.com/).…”

“…Recently, MEC and wMEC approaches have been used in the context of long reads, confirming that long fragments allow to assemble haplotypes more accurately than traditional short reads (Aguiar and Istrail, 2012;Duitama et al, 2012;Patterson et al, 2014Patterson et al, , 2015. Since MEC is NP-hard (Cilibrasi et al, 2007), exact solutions have exponential complexity.…”

“…There were also proposed fixedparameter tractable (FPT) algorithms that take time exponential in the number of variants per read (Bonizzoni et al, 2015;He et al, 2010He et al, , 2013 and, hence, are well-suited for short reads but become unfeasible for long reads. For this kind of data, heuristic approaches have been proposed to respond to the lack of exact solutions (Bansal and Bafna, 2008;Duitama et al, 2012). Most of the proposed heuristics, such as REFHAP (Duitama et al, 2010), make use of the traditional all-heterozygous assumption, that forces the heterozygosity of all the phased positions.…”

“…Most of the proposed heuristics, such as REFHAP (Duitama et al, 2010), make use of the traditional all-heterozygous assumption, that forces the heterozygosity of all the phased positions. These heuristics have good performances but do not offer guarantees on the optimality of the returned solution (Duitama et al, 2012). Two recent articles (Kuleshov, 2014;Patterson et al, 2014) aim at processing future-generation long reads by introducing algorithms exponential in the sequencing coverage, a parameter which is not expected to grow as fast as read length with the advent of future-generation technologies.…”

“…On a real standard benchmark of long reads (Duitama et al, 2012), we executed each tool under the all-heterozygous assumption, since this dataset has low coverage ($3Â on average) and since the covered positions are heterozygous with high confidence. HAPCOL turns out to be competitive with the considered methods, improving the accuracy and the number of phased positions.…”