Optimized double-digest genotyping by sequencing (ddGBS) method with high-density SNP markers and high genotyping accuracy for chickens

Abstract

High-density single nucleotide polymorphism (SNP) markers are crucial to improve the resolution and accuracy of genome-wide association study (GWAS) and genomic selection (GS). Numerous approaches, including whole genome sequencing, genome sampling sequencing, and SNP chips are able to discover or genotype markers at different densities and costs. Achieving an optimal balance between sequencing resolution and budgets, especially in large-scale population genetics research, constitutes a major challenge. Here, we performed improved double-enzyme digestion genotyping by sequencing (ddGBS) on chicken. We evaluated eight double-enzyme digestion combinations, and EcoR I- Mse I was chosen as the optimal combination for the chicken genome. We firstly proposed that two parameters, optimal read-count point (ORP) and saturated read-count point (SRP), could be utilized to determine the optimal sequencing volume. A total of 291,772 high-density SNPs from 824 animals were identified. By validation using the SNP chip, we found that the consistency between ddGBS data and the SNP chip is over 99%. The approach that we developed in chickens, which is high-quality, high-density, cost-effective (300 K, $30/sample), and time-saving (within 48 h), will have broad applications in animal breeding programs.

Double-digest RAD-sequencing: do wet and dry protocol parameters impact biological results?

“…Multiple studies have speculated and tested the impact of different parameter settings on pre- and post-sequencing procedures of RAD-based protocols, especially in loci reconstruction summary statistics, i.e number of loci or SNPs, or heterozygosity (Davey et al, 2013; Gautier et al, 2013; K. Andrews & Luikart, 2014; K. R. Andrews et al, 2014; Puritz et al, 2014; Mastretta-Yanes et al, 2015; Burns et al, 2017; Rochette & Catchen, 2017; Y. Wang et al, 2017; Willis et al, 2017). Nevertheless, only a handful of them linked those parameters to the downstream biological interpretation (Mastretta-Yanes et al, 2015; Rodríguez-Ezpeleta et al, 2016; Shafer et al, 2017; Malinsky et al, 2018).…”

“…Concerning the dry protocol, an important part of the RAD-based sequencing literature pertains to the bioinformatic treatment of sequences and to loci reconstruction (Mastretta-Yanes et al, 2015; Paris et al, 2017; Rochette & Catchen, 2017; Y. Wang et al, 2017). These studies highlight the impact of clustering thresholds ( e.g.…”

“…Indeed, these thresholds have been shown to influence the number of recovered loci, coverage, number of identified SNPs and error rates (Mastretta-Yanes et al, 2015; Paris et al, 2017; Rochette & Catchen, 2017; Y. Wang et al, 2017). In agreement with previous works, we found a substantial impact of the minimum coverage ( m ) and clustering ( M ) thresholds on ddRADseq loci recovery and reconstruction during the bioinformatic process.…”

“…For example, this choice will influence the number of digested fragments, their location in the genome, and their size distribution (Burns et al, 2017; Y. Wang et al, 2017). DNA quality also influences SNPs recovery, because degraded ( i.e.…”

NanoGBS: A Miniaturized Procedure for GBS Library Preparation

“…NGS-based genotyping methods are capable of simultaneously genotyping markers on a genome-wide scale, even in nonmodel species with little or no available genetic information (Torkamaneh et al, 2018a). NGS-based reduced-representation sequencing (RRS) strategies [restriction site-associated DNA sequencing (RAD-seq), complexity reduction of polymorphic sequences (CRoPS), genotyping-bysequencing (GBS), double-digest RAD-seq (ddRAD), 2bRAD, and double-digest GBS (ddGBS)], relying on high-throughput sequencing (HTS) of multiplexed samples, allows for the genotyping of thousands to millions of SNPs in parallel in large sets of individual samples (Baird et al, 2008;Elshire et al, 2011;Davey et al, 2011;Peterson et al, 2012;Wang et al, 2012;Ali et al, 2016;Wang et al, 2017). The RRS methods all follow similar DNA library preparation steps (DNA digestion, adaptor ligation, amplification, and sequencing) and are generically called GBS.…”