An autopolyploid-suitable polyBSA-seq strategy for screening candidate genetic markers linked to leaf blight resistance in sugarcane

Abstract

An autopolyploid-suitable polyBSA-seq strategy was developed for screening candidate genetic markers linked to leaf blight resistance in sugarcane. Due to the complex genome architecture, the quantitative trait loci mappings and linkage marker selections for agronomic traits of autopolyploid crops were mainly limited to the time-consuming and cost intensive construction of genetic maps. To map resistance-linked markers for sugarcane leaf blight (SLB) caused by Stagonospora tainanensis, the autopolyploid-suitable bulk-segregant analysis based on the sequencing (polyBSA-seq) strategy was successfully applied for the first time. Resistant- and susceptible-bulks (R- and S-bulks) constructed from the extreme-phenotypic sugarcane F₁ lines of YT93-159 × ROC22 were deep sequenced with 195.0 × for bulks and 74.4 × for parents. Informative single-dose variants (ISDVs) present as one copy in one parent and null in the other parent were detected based on the genome sequence of LA Purple, an autooctoploid Saccharum officinarum, to screen candidate linkage markers (CLMs). The proportion of the number of short reads harboring ISDVs in the total short reads covering a given genomic position was defined as ISDV index and the ISDVs with indices met the threshold set in this study (0.04-0.14) were selected as CLMs. In total, three resistance- and one susceptibility-related CLMs for SLB resistance were identified by the polyBSA-seq. Among them, two markers on chromosome 10 were less than 300 Kb apart. Furthermore, the RNA-seq was used to calculate the expression level of genes within 1.0 Mb from the aforementioned four CLMs, which demonstrated that twelve genes were differentially expressed between resistant and susceptible clones, including a receptor-like kinase and an ethylene-responsive transcription factor. This is the first reported polyBSA-seq in autopolyploid sugarcane, which specifically tailored for the fast selection of the CLMs and causal genes associated with important agronomic traits.