Double Digest RADseq: An Inexpensive Method for De Novo SNP Discovery and Genotyping in Model and Non-Model Species

By Brant Peterson, Jesse N. Weber, Emily H. Kay, H. Fisher, H. Hoekstra. PLoS ONE, 2012

3,452 citations recorded by the source corpus.

The ability to efficiently and accurately determine genotypes is a keystone technology in modern genetics, crucial to studies ranging from clinical diagnostics, to genotype-phenotype association, to reconstruction of ancestry and the detection of selection. To date, high capacity, low cost genotyping has been largely achieved via “SNP chip” microarray-based platforms which require substantial prior knowledge of both genome sequence and variability, and once designed are suitable only for those targeted variable nucleotide sites. This method introduces substantial ascertainment bias and inherently precludes detection of rare or population-specific variants, a major source of information for both population history and genotype-phenotype association. Recent developments in reduced-representation genome sequencing experiments on massively parallel sequencers (commonly referred to as RAD-tag or RADseq) have brought direct sequencing to the problem of population genotyping, but increased cost and procedural and analytical complexity have limited their widespread adoption. Here, we describe a complete laboratory protocol, including a custom combinatorial indexing method, and accompanying software tools to facilitate genotyping across large numbers (hundreds or more) of individuals for a range of markers (hundreds to hundreds of thousands). Our method requires no prior genomic knowledge and achieves per-site and per-individual costs below that of current SNP chip technology, while requiring similar hands-on time investment, comparable amounts of input DNA, and downstream analysis times on the order of hours. Finally, we provide empirical results from the application of this method to both genotyping in a laboratory cross and in wild populations. Because of its flexibility, this modified RADseq approach promises to be applicable to a diversity of biological questions in a wide range of organisms.

Subfields: Molecular Biology

View Paper: https://www.semanticscholar.org/paper/b0114e2eb305bd7c12d47b03e8c57964a1a41f22

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Citations in the corpus

Genotyping-By-Sequencing for Plant Genetic Diversity Analysis: A Lab Guide for SNP Genotyping (2014, neutral): “[13], each containing priming and ligating sites.”
High-throughput linkage mapping of Australian white cypress pine (Callitris glaucophylla) and map transferability to related species (2015, methodology): “A recently developed RAD sequencing (restriction site associated DNA sequencing) is an efficient technique to obtain SNP genotype data of population samples even for species with no prior genomic knowledge (Baird et al. 2008; Peterson et al. 2012).”
BA3‐SNPs: Contemporary migration reconfigured in BayesAss for next‐generation sequence data (2019, methodology): “Performance benchmarking was conducted for the above programs by analysing 618 files containing empirical data produced via ddRAD sequencing methods (Peterson et al., 2012).”
Research Priorities to Support Effective Manta and Devil Ray Conservation (2018, methodology): “, 2012), we recommend high-throughput genomic methods (Peterson et al., 2012; Catchen et al., 2013) applied to samples of representative coverage within and among populations to generate robust estimates of population structure.”
Marker-Assisted Plant Breeding: Principles and Practices (2015, neutral): “Further, its graph clustering procedure can group haplotypes with any number of mismatches (Peterson et al. 2012).”
ipyrad: Interactive assembly and analysis of RADseq datasets (2020, background): “…(McKain et al., 2018), restriction site-associated DNA sequencing (RADseq) and related methods have become a popular choice for their flexibility and affordability (Andrews et al., 2016; Baird et al., 2008; Davey et al., 2011; Elshire et al., 2011; Miller et al., 2007; Peterson et al., 2012).”
Short Tree, Long Tree, Right Tree, Wrong Tree: New Acquisition Bias Corrections for Inferring SNP Phylogenies (2015, methodology): “…site associated DNA sequencing (RADseq) has become a popular method for generating single nucleotide polymorphism (SNP) data sets in non-model organisms, because the method requires little to no prior knowledge of the genome (Baird et al. 2008; Seeb et al. 2011; Peterson et al. 2012).”
High-Throughput Genomic Data in Systematics and Phylogenetics (2013, methodology): “Other limitations of RAD-seq are reviewed elsewhere (Davey et al. 2012, Arnold et al. 2013).”
Special features of RAD Sequencing data: implications for genotyping (2012, methodology): “This suggests it would be valuable to develop alternative methods of shearing restriction fragments, perhaps using enzymatic protocols such as Nextera or dsDNA fragmentase (Adey et al. 2010), or by using additional restriction enzymes (Peterson et al .”
Species Delimitation using Genome-Wide SNP Data (2013, methodology): “To collect genome-wide sampling of SNPs, we followed the laboratory protocols for double-digest RADseq (ddRADseq) described in Peterson et al. (2012).”
A framework for incorporating evolutionary genomics into biodiversity conservation and management (2015, neutral): “See [109-111].”
Using nuclear gene data for plant phylogenetics: Progress and prospects II. Next‐gen approaches (2015, methodology): “…have been used recently for studies of plant (Scaglione et al., 2012; Eaton & Ree, 2013; Wang et al., 2013a, 2013b; Hipp et al., 2014) and animal groups (Emerson et al., 2010; Peterson et al., 2012; Reitzel et al., 2013; Wagner et al., 2013; Cruaud et al., 2014; Takahashi et al., 2014).”
dartr: An r package to facilitate analysis of SNP data generated from reduced representation genome sequencing (2018, neutral): “…sequencing (RADSeq), double digest RADSeq (ddRADSeq) and DarTSeq (Andrews, Good, Miller, Luikart, & Hohenlohe, 2016; Baird et al., 2008; Jaccoud, Peng, Feinstein, & Kilian, 2001; Kilian et al., 2012; Peterson, Weber, Kay, Fisher, & Hoekstra, 2012; Sansaloni et al., 2011; van Tassell et al., 2008).”
Harnessing the power of RADseq for ecological and evolutionary genomics (2016, methodology): “; double digest RAD (ddRAD), data from Table 1 in Peterson et al.(17) and allowing for up to double the size range; ezRAD, data from Toonen et al.”
vcfr: a package to manipulate and visualize variant call format data in R (2017, background): “2011), double-digest RAD-seq (ddRAD) (Peterson et al. 2012) and amplicon sequencing (GT-Seq) (Campbell et al.”