Linkage mapping and QTL analysis of flowering time using ddRAD sequencing with genotype error correction in Brassica nap
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RESEARCH ARTICLE
Open Access
Linkage mapping and QTL analysis of flowering time using ddRAD sequencing with genotype error correction in Brassica napus Armin Scheben1,2, Anita A. Severn-Ellis1, Dhwani Patel1, Aneeta Pradhan1, Stephen J. Rae3, Jacqueline Batley1 and David Edwards1*
Abstract Background: Brassica napus is an important oilseed crop cultivated worldwide. During domestication and breeding of B. napus, flowering time has been a target of selection because of its substantial impact on yield. Here we use double digest restriction-site associated DNA sequencing (ddRAD) to investigate the genetic basis of flowering in B. napus. An F2 mapping population was derived from a cross between an early-flowering spring type and a lateflowering winter type. Results: Flowering time in the mapping population differed by up to 25 days between individuals. High genotype error rates persisted after initial quality controls, as suggested by a genotype discordance of ~ 12% between biological sequencing replicates. After genotype error correction, a linkage map spanning 3981.31 cM and compromising 14,630 single nucleotide polymorphisms (SNPs) was constructed. A quantitative trait locus (QTL) on chromosome C2 was detected, covering eight flowering time genes including FLC. Conclusions: These findings demonstrate the effectiveness of the ddRAD approach to sample the B. napus genome. Our results also suggest that ddRAD genotype error rates can be higher than expected in F2 populations. Quality filtering and genotype correction and imputation can substantially reduce these error rates and allow effective linkage mapping and QTL analysis. Keywords: Brassica napus, Double digest restriction-site associated DNA sequencing, Flowering, QTL, SNP
Background Genotyping-by-sequencing (GBS) is a powerful tool for high-throughput discovery of genetic polymorphisms in crops [1–5]. GBS comprises a range of library preparation and sequencing approaches that differ in their costs, methodical biases and the type and amount of data produced [1, 6]. Restriction site-associated DNA * Correspondence: [email protected] 1 School of Biological Sciences and Institute of Agriculture, The University of Western Australia, Perth, WA, Australia Full list of author information is available at the end of the article
sequencing (RAD) is a GBS method that can be used to cost-effectively calibrate the number and coverage of genotyped loci and single nucleotide polymorphisms (SNPs) by varying the enzymes used and the sequencing depth. A recent comparative analysis of single enzyme RAD and two enzyme double digest RAD (ddRAD) used a range of enzyme combinations in different plants and suggested that the enzyme combination of HinfI and HpyCH4IV was promising for maximising genome coverage breadth across a range of species [7]. Like other GBS approaches, ddRAD is prone to missing data and
© The Author(s). 2020 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distrib
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