Conversion of array‐based single nucleotide polymorphic markers for use in targeted genotyping by sequencing in hexaploid wheat (Triticum aestivum)

Abstract

SummaryWheat breeders and academics alike use single nucleotide polymorphisms (SNPs) as molecular markers to characterize regions of interest within the hexaploid wheat genome. A number of SNP‐based genotyping platforms are available, and their utility depends upon factors such as the available technologies, number of data points required, budgets and the technical expertise required. Unfortunately, markers can rarely be exchanged between existing and newly developed platforms, meaning that previously generated data cannot be compared, or combined, with more recently generated data sets. We predict that genotyping by sequencing will become the predominant genotyping technology within the next 5–10 years. With this in mind, to ensure that data generated from current genotyping platforms continues to be of use, we have designed and utilized SNP‐based capture probes from several thousand existing and publicly available probes from Axiom® and KASP™ genotyping platforms. We have validated our capture probes in a targeted genotyping by sequencing protocol using 31 previously genotyped UK elite hexaploid wheat accessions. Data comparisons between targeted genotyping by sequencing, Axiom® array genotyping and KASP™ genotyping assays, identified a set of 3256 probes which reliably bring together targeted genotyping by sequencing data with the previously available marker data set. As such, these probes are likely to be of considerable value to the wheat community. The probe details, full probe sequences and a custom built analysis pipeline may be freely downloaded from the CerealsDB website (http://www.cerealsdb.uk.net/cerealgenomics/CerealsDB/sequence_capture.php).

Analysis of the recombination landscape of hexaploid bread wheat reveals genes controlling recombination and gene conversion frequency

“…We could directly associate on average 95.7% of our previously defined GCs with GC events in the sequencing data that were on average 29,992,400 bp in length with 61.2% of events < 20 Mbp (Table 1). Furthermore, on average, we could validate 83.0% of the specific array SNP alleles that were used to define GCs and given the reported agreement rates of 85.7% for SNP arrays when compared to sequencing data by Burridge et al [5]; this is in line with expectations. Using the array, the reason for our ability to identify GCs in low-resolution genotyping data is that we typically detect the larger GC events whereas whole genome sequencing gives the ability to robustly detect shorter GC events across the genome.…”

Analysis of the recombination landscape of hexaploid bread wheat reveals genes controlling recombination and gene conversion frequency

“…We could directly associate on average 95.7% of our previously defined GCs with GC events in the sequencing data that were on average 29,992,400bp in length with 61.2% of events <20Mbp ( Table 2). Furthermore on average we could validate 83.0% of the specific array SNP alleles that were used to define GCs and given the reported agreement rates of 85.7% for SNP arrays when compared to sequencing data by Burridge et al (2017), this is in line with expectations. Using the array, the reason for our ability to identify GCs in low-resolution genotyping data is that we typically detect the larger GC events whereas whole genome sequencing gives the ability to robustly detect shorter GC events across the genome.…”

Developing a High-Throughput SNP-Based Marker System to Facilitate the Introgression of Traits From Aegilops Species Into Bread Wheat (Triticum aestivum)

“…Leaf tissue was harvested 4 weeks after germination, frozen in liquid nitrogen and stored at −20°C prior to nucleic acid extraction. Genomic DNA was prepared using a phenol–chloroform extraction method (Burridge et al, 2017), treated with RNase-A (QIAGEN Ltd., Manchester, UK) according to the manufacturer's instructions and purified using the QiaQuick PCR purification kit (QIAGEN Ltd).…”