Analysis of the Rpv12 locus in a haplotype‑separated grapevine genome sequence

Plasmopara viticola, the grapevine downy mildew pathogen, causes severe losses in viticulture if not counteracted by fungicide sprays that need to be repeatedly applied during each growing season. To reduce the amount of plant protection, modern grapevine breeding generates fungus‑resistant grapevine cultivars by introgression of resistance loci from wild Vitis spec. sources. However, the presence of only a single resistance locus may provoke the emergence of pathogen races able to overcome the resistance trait of the host. Therefore, a combination of several, independently acting resistance loci is required for sustainable genetic resistance. Quite little is known about the resistance‑conferring genes within the various grapevine resistance loci. To ameliorate this situation and make stacking of resistance loci more efficient, the Rpv12 locus originating from the Asian Vitis amurensis was sequenced and characterized. The complete genome of breeding line Gf.99‑03, carrying Rpv12 in heterozygous state, was analyzed. Haplotypes were resolved by assigning the reads to one of the parents of Gf.99‑03 using trio binning. Annotation of the resulting genomic sequences was based on RNA-Seq data and predicted gene models. The haplotype carrying the Rpv12 locus, delimited by markers UDV‑014 and UDV‑370 on chromosome 14 (Venuti et al., 2013), diverges strongly from the susceptible haplotype as well as from the reference genome PN40024 12X.v2. It was found to contain two important gene clusters. One cluster includes pathogen-inducible genes similar to the gene ACCELERATED CELL DEATH 6 (A. thaliana) likely involved in hypersensitive response upon pathogen attack. The second cluster comprises positional resistance candidate genes corresponding to typical NLRs (nucleotide binding site, leucine rich repeats), hypothesized to be involved in pathogen perception and cellular defense signalling.