Hybridization Capture Using RAD Probes (hyRAD), a New Tool for Performing Genomic Analyses on Collection Specimens

Abstract

In the recent years, many protocols aimed at reproducibly sequencing reduced-genome subsets in non-model organisms have been published. Among them, RAD-sequencing is one of the most widely used. It relies on digesting DNA with specific restriction enzymes and performing size selection on the resulting fragments. Despite its acknowledged utility, this method is of limited use with degraded DNA samples, such as those isolated from museum specimens, as these samples are less likely to harbor fragments long enough to comprise two restriction sites making possible ligation of the adapter sequences (in the case of double-digest RAD) or performing size selection of the resulting fragments (in the case of single-digest RAD). Here, we address these limitations by presenting a novel method called hybridization RAD (hyRAD). In this approach, biotinylated RAD fragments, covering a random fraction of the genome, are used as baits for capturing homologous fragments from genomic shotgun sequencing libraries. This simple and cost-effective approach allows sequencing of orthologous loci even from highly degraded DNA samples, opening new avenues of research in the field of museum genomics. Not relying on the restriction site presence, it improves among-sample loci coverage. In a trial study, hyRAD allowed us to obtain a large set of orthologous loci from fresh and museum samples from a non-model butterfly species, with a high proportion of single nucleotide polymorphisms present in all eight analyzed specimens, including 58-year-old museum samples. The utility of the method was further validated using 49 museum and fresh samples of a Palearctic grasshopper species for which the spatial genetic structure was previously assessed using mtDNA amplicons. The application of the method is eventually discussed in a wider context. As it does not rely on the restriction site presence, it is therefore not sensitive to among-sample loci polymorphisms in the restriction sites that usually causes loci dropout. This should enable the application of hyRAD to analyses at broader evolutionary scales.

Spatial and temporal genetic dynamics of the grasshopper Oedaleus decorus revealed by museum genomics

“…We prepared Illumina libraries following the recently developed hyRAD protocol, which allows us to sequence a representative fraction of the genome in a set of samples (Suchan et al, 2016). This method uses hybridization capture and does not strictly rely on PCR amplification.…”

“…Here, we combined a classical shotgun protocol with hybridization capture of restriction site-associated DNA loci (hyRAD)-a recently developed technique using fragments generated by RAD as baits to capture homologous sequences generated by shotgun and easily applicable to any nonmodel species (Suchan et al, 2016)-to investigate the spatial and temporal genetic dynamics of the endangered grasshopper Oedaleus decorus (Acrididae, Germar 1826). The species is characterized by a large South-Palearctic distribution, ranging from Macaronesian Islands through the Mediterranean Basin to central Asia (Schmidt & Lilge, 1997).…”

Comparing Pool‐seq, Rapture, and GBS genotyping for inferring weak population structure: The American lobster (Homarus americanus) as a case study

“…For the past few years, sequence capture approaches have undergone a new upswing, in particular with the concept of targeted sequence enrichment that couples the power of sequence capture with NGS technology (Grover, Salmon, & Wendel, 2012). More recently, methods combining sequence capture enrichment and reduced representation libraries have been proposed (Ali et al, 2016;Boucher et al, 2016;Hoffberg et al, 2016;Suchan et al, 2016).…”

Molecular ecology studies of species radiations: current research gaps, opportunities and challenges

“…4B), WGS and sequence capture may be best suited for addressing the current major research gaps and challenges faced by studies of species radiations, which can be summarized concisely as follows: (i) to tackle the many 'nonmodel' organismal groups currently underrepresented in studies of radiations (Fig. 2), including those for which high-quality tissue samples are difficult to come by or specimens from herbaria or museum collections (Suchan et al 2016), (ii) to scale up the sampling of populations and species in order to move the field forward from 'speciation genomics' towards 'evolutionary radiation genomics', thus filling the sparsely populated upper right portion of sample size space exemplified in Fig. 3, (iii) to achieve more elaborate and representative sampling of nuclear genomes in studies of radiations, (iv) to increasingly capture those genes and regulatory elements that matter most to the evolutionary process, in addition to neutrally evolving sequence regions.…”