Mapping‐by‐sequencing in complex polyploid genomes using genic sequence capture: a case study to map yellow rust resistance in hexaploid wheat

Abstract

SummaryPreviously we extended the utility of mapping‐by‐sequencing by combining it with sequence capture and mapping sequence data to pseudo‐chromosomes that were organized using wheat–Brachypodium synteny. This, with a bespoke haplotyping algorithm, enabled us to map the flowering time locus in the diploid wheat Triticum monococcum L. identifying a set of deleted genes (Gardiner et al., 2014). Here, we develop this combination of gene enrichment and sliding window mapping‐by‐synteny analysis to map the Yr6 locus for yellow stripe rust resistance in hexaploid wheat. A 110 MB NimbleGen capture probe set was used to enrich and sequence a doubled haploid mapping population of hexaploid wheat derived from an Avalon and Cadenza cross. The Yr6 locus was identified by mapping to the POPSEQ chromosomal pseudomolecules using a bespoke pipeline and algorithm (Chapman et al., 2015). Furthermore the same locus was identified using newly developed pseudo‐chromosome sequences as a mapping reference that are based on the genic sequence used for sequence enrichment. The pseudo‐chromosomes allow us to demonstrate the application of mapping‐by‐sequencing to even poorly defined polyploidy genomes where chromosomes are incomplete and sub‐genome assemblies are collapsed. This analysis uniquely enabled us to: compare wheat genome annotations; identify the Yr6 locus – defining a smaller genic region than was previously possible; associate the interval with one wheat sub‐genome and increase the density of SNP markers associated. Finally, we built the pipeline in iPlant, making it a user‐friendly community resource for phenotype mapping.

Applying the latest advances in genomics and phenomics for trait discovery in polyploid wheat

“…Instead, integrating known genetic mapping with new genomic approaches can accelerate trait discovery. Exome‐capture sequencing has been used to identify variation in the coding region of the wheat genome (Saintenac et al ., ) and can be used to identify candidate genes, often in combination with bulk‐segregant analysis (Gardiner et al ., ; Mo et al ., ). SNPs identified from the exome capture can be used to both develop new markers for genetic mapping and to identify mutations in putative candidate genes.…”

Integrating genomic resources to present full gene and promoter capture probe sets for bread wheat

“…To date, such diversity has been profiled in wheat using capture probe sets that have not been able to make use of the recent advances in wheat genome sequencing and annotation. Most of the diversity studies have implemented either cDNA/exon-based probe sets of 56 and 84 Mb (Winfield et al, 2012;Krasileva et al, 2016) or the gene-based probe set of 107 Mb (Jordan et al, 2015;Gardiner et al, 2016). Aligning the 107 Mb capture probe set to the current wheat genome annotations (BLASTN, e-value 1e-05, identity 95%, minimum length 40bp), we can see that it represents only 32.9% of the high confidence gene-set or 21.2% of the gene-set plus promoters defined as 2Kbp upstream (Clavijo et al, 2017).…”

Adding Complexity to Complexity: Gene Family Evolution in Polyploids

“…We conclude with recommendations for how some of these challenges might be overcome using deep sequencing approaches. We reiterate the recommendation from others (Salmon et al, 2012;Jupe et al, 2013;Gardiner et al, 2016;Van Weymers et al, 2016;Krasileva et al, 2017) that non-amplicon based targeted sequence capture (e.g., whole genome exon capture or targeting of particular gene families) is the most promising method for tackling the full complexity of gene family evolution in complex genomes but suggest cautionary strategies that should be considered when interpreting evolutionary patterns.…”

“…Fortunately, rapid advances in sequencing technology and bioinformatic processing mean that the toolbox available to resolve such challenges continues to improve. Targeted sequence capture, for example, has been used effectively to investigate genomic changes in polyploids (Salmon et al, 2012;Gardiner et al, 2016;Krasileva et al, 2017). However, even with these advances in technology there are important issues to consider when resolving and interpreting evolutionary dynamics of gene families, particularly for systems in which recent polyploidization and hybridization could complicate accurate assembly into orthologs and subsequent genotyping within and between copies.…”