Genome Wide Association Studies in Multiple Spinach Breeding Populations Refine Downy Mildew Race 13 Resistance Genes

Abstract

Downy mildew, caused by the oomycete Peronospora effusa, is the most economically important disease on spinach. Fourteen new races of P. effusa have been identified in the last three decades. The frequent emergence of new races of P. effusa continually overcome the genetic resistance to the pathogen. The objectives of this research were to more clearly map the downy mildew resistance locus RPF1 in spinach, to identify single nucleotide polymorphism (SNP) markers associated with the resistance, and to refine the candidate genes responsible for the resistance. Progeny from populations generated from crosses of cultivars resistant (due to RPF1) to race 13 of P. effusa (Swan, T-Bird, Squirrel, and Tonga) with race 13 susceptible cultivars (Whale and Polka) were inoculated and the downy mildew disease response determined. Association analysis was performed in TASSEL, GAPIT, PLINK, and GENESIS programs using SNP markers identified from genotyping by sequencing (GBS). Association analysis mapped the race 13 resistance loci (RPF1) to positions 0.39, 0.69, 0.94-0.98, and 1.2 Mb of chromosome 3. The associated SNPs were within 1-7 kb of the disease resistance genes Spo12784, Spo12719, Spo12905, and Spo12821, and 11-18 Kb from Spo12903. This study extended our understanding of the genetic basis of downy mildew resistance in spinach and provided the most promising candidate genes Spo12784 and Spo12903 near the RPF1 locus, to pursue functional validation. The SNP markers may be used to select for the resistant lines to improve genetic resistance against the downy mildew pathogen and in developing durably resistant cultivars.

Genomic analyses provide insights into spinach domestication and the genetic basis of agronomic traits

“…We identified a region on chromosome 3 showing strong LD and significant association with DM resistance. This region overlaps with genomic interval identified in previous study 45 and encodes several R genes with one being promising candidate for future functional validation. In addition to the major association signal, minor signals were identified on chromosome 4 and 5, which have not been reported before, suggesting the diversity of DM resistance in the spinach population and providing a great resource for spinach breeding.…”

Genotyping by sequencing suggests overwintering of Peronospora destructor in southwestern Québec, Canada

“…humuli , known to have both clonal and homothallic reproduction modes (Gent et al, 2017, 2019). Such high linkage disequilibrium may introduce redundancy, especially for large genomic data from populations with small effective sample size, and this may limit the resolution in identifying informative loci (Bhattarai et al, 2020; Calus & Vandenplas, 2018). In our study, most of the variant loci had G ′ ST values close to 0 and were randomly distributed throughout the genome even after heavy data pruning, which is characteristic of random sequencing errors and abiotic variation sources (Gent et al, 2019).…”

“…Typically, downy mildews produce numerous wind‐dispersed sporangia (asexual spores), spreading the inoculum over short or long distances during the growing season. It is generally accepted that downy mildew pathogen populations are mostly clonal (Bhattarai et al, 2020; Blanco‐Meneses et al, 2018; Gent et al, 2019; Lyon et al, 2016; Summers, Gulliford, et al, 2015). However, they are also capable of genetic recombination through sexual reproduction (homothallism and heterothallism), producing oospores, which are long‐term survival structures that can, like Peronospora destructor , survive up to 25 years in soils (McKay, 1957).…”

Genetic Diversity, Structure, and Selective Sweeps in Spinacia turkestanica Associated With the Domestication of Cultivated Spinach

“…The DNA quality was checked on a 1% agarose gel, quantified using a Qubit, and submitted to the University of Wisconsin Madison Biotechnology Center ( https://www.biotech.wisc.edu/ ) for sequencing, where DNA quality and integrity were re-evaluated using the Quant-IT PicoGreen fluorescent dye (Thermo Fisher, Waltham, MA). The GBS method of Elshire et al (2011) was used to sequence the samples after digesting genomic DNA with the Ape KI restriction enzyme, as described by Bhattarai et al (2020) . Digested DNA fragments were ligated with unique barcodes and Illumina adapters, and the samples were pooled in equal proportion to construct GBS libraries, as described by Elshire et al (2011) .…”