RNA‐Seq bulked segregant analysis enables the identification of high‐resolution genetic markers for breeding in hexaploid wheat. Issue 5 (8th November 2014)

• Record Type: Journal Article
• Authors: Ramirez‐Gonzalez, Ricardo H.; Segovia, Vanesa; Bird, Nicholas; Fenwick, Paul; Holdgate, Sarah; Berry, Simon; Jack, Peter; Caccamo, Mario; Uauy, Cristobal
• Is Part Of: Plant biotechnology journal. Volume 13:Issue 5(2015:Jun.)
• Page Start: 613
• Page End: 624
• Publication Date: 2014-11-08
• Subjects: Plant biotechnology -- Periodicals; Plant genetic engineering -- Periodicals; 630.272
• DOI: 10.1111/pbi.12281
• Languages: English
• ISSNs: 1467-7644

Abstract

<abstract abstract-type="main" id="pbi12281-abs-0001"> <title>Summary</title> <p>The identification of genetic markers linked to genes of agronomic importance is a major aim of crop research and breeding programmes. Here, we identify markers for <italic>Yr15</italic>, a major disease resistance gene for wheat yellow rust, using a segregating F<sub>2</sub> population. After phenotyping, we implemented RNA sequencing (RNA‐Seq) of bulked pools to identify single‐nucleotide polymorphisms (SNP) associated with <italic>Yr15</italic>. Over 27 000 genes with SNPs were identified between the parents, and then classified based on the results from the sequenced bulks. We calculated the bulk frequency ratio (BFR) of SNPs between resistant and susceptible bulks, selecting those showing sixfold enrichment/depletion in the corresponding bulks (BFR > 6). Using additional filtering criteria, we reduced the number of genes with a putative SNP to 175. The 35 SNPs with the highest BFR values were converted into genome‐specific KASP assays using an automated bioinformatics pipeline (PolyMarker) which circumvents the limitations associated with the polyploid wheat genome. Twenty‐eight assays were polymorphic of which 22 (63%) mapped in the same linkage group as <italic>Yr15</italic>. Using these markers, we mapped <italic>Yr15</italic> to a 0.77‐cM interval. The three most closely linked SNPs were tested across varieties and breeding lines representing UK elite germplasm. Two flanking markers were diagnostic in over 99% of lines tested, thus providing a reliable haplotype for marker‐assisted selection in these breeding programmes. Our results demonstrate that the proposed methodology can be applied in polyploid F<sub>2</sub> populations to generate high‐resolution genetic maps across target intervals.</p> </abstract>