High-resolution bulked segregant analysis enables candidate gene identification for bacterial wilt resistance in Italian ryegrass

Abstract

AbstractBacterial wilt, caused byXanthomonas translucenspv.graminis(Xtg), is a serious disease of economically important forage grasses, including Italian ryegrass (Lolium multiflorumLam.). A major QTL for resistance toXtgwas previously identified, but the precise location as well as the genetic factors underlying the resistance are yet to be determined. To this end, we applied a bulked segregant analysis (BSA) approach, using whole-genome deep sequencing of pools of the most resistant and most susceptible individuals of a large (n = 7,484) biparental F2population segregating for resistance toXtg. Using chromosome-level genome sequences as references, we were able to define a ∼300 kb region highly associated to resistance on pseudo-chromosome 4. Further investigation of this region revealed multiple genes with a known role in disease resistance, including genes encoding for Pik2-like disease resistance proteins, cysteine-rich kinases, and RGA4- and RGA5-like disease resistance proteins. Investigation of allele frequencies in the pools and comparative genome analysis in the grandparents of the F2population revealed that some of these genes contain variants with allele frequencies that correspond to the expected heterozygosity in the resistant grandparent. This study emphasizes the efficacy combining BSA studies in very large populations with whole genome deep sequencing and high-quality genome sequences to pinpoint regions associated with a binary trait of interest and accurately define a small set of candidate genes. Furthermore, markers identified in this region hold significant potential for marker-assisted breeding strategies to breed resistance toXtgin Italian ryegrass cultivars more efficiently.