High-throughput genotyping by whole-genome resequencing

TLDR

Next‑generation sequencing and the growing number of genome sequences open opportunities to redesign genotyping strategies for more effective genetic mapping and genome analysis, and this genome‑based method may replace conventional marker‑based genotyping to provide a powerful tool for large‑scale gene discovery and addressing a wide range of biological questions. The authors developed a high‑throughput method for genotyping recombinant populations using whole‑genome resequencing data generated by the Illumina Genome Analyzer. The method employs a sliding‑window approach to jointly analyze genome‑wide SNPs from Illumina resequencing data for genotype calling and recombination breakpoint determination. The approach produced a genetic map for 150 rice recombinant inbred lines with 99.94 % genotype accuracy and ~40 kb breakpoint resolution, was ~20× faster and ~35× more precise than a 287‑marker PCR map, identified a large‑effect QTL for plant height, and proved robust across diverse mapping populations and genome sizes.

Abstract

The next-generation sequencing technology coupled with the growing number of genome sequences opens the opportunity to redesign genotyping strategies for more effective genetic mapping and genome analysis. We have developed a high-throughput method for genotyping recombinant populations utilizing whole-genome resequencing data generated by the Illumina Genome Analyzer. A sliding window approach is designed to collectively examine genome-wide single nucleotide polymorphisms for genotype calling and recombination breakpoint determination. Using this method, we constructed a genetic map for 150 rice recombinant inbred lines with an expected genotype calling accuracy of 99.94% and a resolution of recombination breakpoints within an average of 40 kb. In comparison to the genetic map constructed with 287 PCR-based markers for the rice population, the sequencing-based method was ∼20× faster in data collection and 35× more precise in recombination breakpoint determination. Using the sequencing-based genetic map, we located a quantitative trait locus of large effect on plant height in a 100-kb region containing the rice “green revolution” gene. Through computer simulation, we demonstrate that the method is robust for different types of mapping populations derived from organisms with variable quality of genome sequences and is feasible for organisms with large genome sizes and low polymorphisms. With continuous advances in sequencing technologies, this genome-based method may replace the conventional marker-based genotyping approach to provide a powerful tool for large-scale gene discovery and for addressing a wide range of biological questions.