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The genome sequence of the diploid and highly homozygous Vitis vinifera genotype PN40024 serves as the reference for many grapevine studies. Despite several improvements to the PN40024 genome assembly, its current version PN12X.v2 is quite fragmented and only represents the haploid state of the genome with mixed haplotypes. In fact, being nearly homozygous, this genome contains several heterozygous regions that are yet to be resolved. Taking the opportunity of improvements that long-read sequencing technologies offer to fully discriminate haplotype sequences, an improved version of the reference, called PN40024.v4, was generated. Through incorporating long genomic sequencing reads to the assembly, the continuity of the 12X.v2 scaffolds was highly increased with a total number decreasing from 2,059 to 640 and a reduction in N bases of 88%. Additionally, the full alternative haplotype sequence was built for the first time, the chromosome anchoring was improved and the number of unplaced scaffolds was reduced by half. To obtain a high-quality gene annotation that outperforms previous versions, a liftover approach was complemented with an optimized annotation workflow for Vitis. Integration of the gene reference catalogue and its manual curation have also assisted in improving the annotation, while defining the most reliable estimation of 35,230 genes to date. Finally, we demonstrated that PN40024 resulted from 9 selfings of cv. “Helfensteiner” (cross of cv. “Pinot noir” and “Schiava grossa”) instead of a single “Pinot noir”. These advances will help maintain the PN40024 genome as a gold-standard reference, also contributing toward the eventual elaboration of the grapevine pangenome.

The downy mildew disease caused by the oomycete Plasmopara viticola is a serious threat for grapevine and can cause enormous yield losses in viticulture. The quantitative trait locus Rpv12 , mediating resistance against P. viticola , was originally found in Asian Vitis amurensis . This locus and its genes were analyzed here in detail. A haplotype-separated genome sequence of the diploid Rpv12 -carrier Gf.99-03 was created and annotated. The defense response against P. viticola was investigated in an infection time-course RNA-seq experiment, revealing approximately 600 upregulated Vitis genes during host–pathogen interaction. The Rpv12 regions of the resistance and the sensitivity encoding Gf.99-03 haplotype were structurally and functionally compared with each other. Two different clusters of resistance-related genes were identified within the Rpv12 locus. One cluster carries a set of four differentially expressed genes with three ACCELERATED CELL DEATH 6-like genes. The other cluster carries a set of six resistance gene analogs related to qualitative pathogen resistance. The Rpv12 locus and its candidate genes for P. viticola resistance provide a precious genetic resource for P. viticola resistance breeding. Newly developed co-segregating simple sequence repeat markers in close proximity to the R -genes enable its improved applicability in marker-assisted grapevine breeding.

The downy mildew disease caused by the oomycete Plasmopara viticola is a serious threat for grapevine and can cause enormous yield losses in viticulture. The quantitative trait locus Rpv12, mediating resistance against P. viticola, was originally found in Asian Vitis amurensis. This locus and its genes were analyzed here in detail. A haplotype-separated genome sequence of the diploid Rpv12-carrier Gf.99-03 was created and annotated. The defense response against P. viticola was investigated in an infection time-course RNA-Seq experiment, revealing approximately 600 up-regulated Vitis genes during host-pathogen interaction. The Rpv12 regions of the resistance conferring and the sensitivity encoding Gf.99-03 haplotypes were structurally and functionally compared to each other. Two different clusters of resistance-related genes were identified within the Rpv12 locus. One cluster carries a set of four differentially expressed genes with three ACCELERATED CELL DEATH 6-like genes. The other cluster carries a set of six resistance gene analogues related to qualitative pathogen resistance. The Rpv12 locus and its candidate genes for P. viticola resistance provide a precious genetic resource for P. viticola resistance breeding. Newly developed co-segregating simple sequence repeat markers in close proximity to the R-genes enable its improved applicability in marker-assisted grapevine breeding.

The downy mildew disease caused by the oomycete Plasmopara viticola is a serious threat for grapevine and can cause enormous yield losses in viticulture. The quantitative trait locus Rpv12, mediating resistance against P. viticola, was originally found in the Asian Vitis amurensis species. Here, we present a detailed analysis of this locus and its genes. The Rpv12-carrying breeding line Gf.99-03 was long-read sequenced and a high-quality haplotype separated genome sequence assembly including gene annotation was worked out. The defense response against P. viticola was investigated through an infection time-course experiment. Differential gene expression analysis revealed approximately 600 up-regulated genes during infection. The Rpv12 region of both haplotypes of Gf.99-03, one conferring resistance and the other one sensitivity, were structurally and functionally compared to each other as well as to the corresponding alleles of the susceptible grapevine model cultivar PN40024 and revealed great structural differences. Two diverse clusters with important resistance related genes were identified that clearly characterize the Rpv12 resistance locus. One cluster carries a set of four differentially expressed genes with three ACCELERATED CELL DEATH 6 related genes. The other cluster carries a set of six resistance gene analogues for potential qualitative pathogen resistance. Co-segregating simple sequence repeat markers for Rpv12 are located in close proximity to the R-genes. The Rpv12 locus and its candidate genes for P. viticola resistance provide a precious genetic resource for P. viticola resistance breeding. Competing Interest Statement The authors have declared no competing interest. Footnotes * Removal of nonfunctional links, update of literature in supplemental methods

The phylloxera resistant rootstock cultivar Boerner is an interspecific hybrid derived from Vitis riparia and V. cinerea and a valuable resource for Vitis disease resistances. We created a fully phased, high-quality Boerner genome sequence named BoeRC using long PacBio reads. Comprehensive gene annotation of both Boerner haplotypes, desig-nated BoeRip and BoeCin, was applied to describe the phylloxera resistance locus Rdv1. Using a mapping population derived from a susceptible V. vinifera breeding line and Boerner, the Rdv1 locus was further delimited. Rdv1, which is derived from V. cinerea and included in the haplotype BoeCin, was compared with sequences of phylloxera-susceptible and phylloxera-tolerant cultivars. Between flanking regions that display high synteny, we detected and precisely characterized a diverse sequence region that covers between 202 to 403 kbp in different haplotypes. In BoeCin, five putative disease re-sistance genes were identified that represent likely candidates for conferring resistance to phylloxera. Competing Interest Statement The authors have declared no competing interest. Footnotes * Improved wording, updated supplements * https://doi.org/10.4119/unibi/2962639 * https://doi.org/10.4119/unibi/2962793

Genomic long reads of the interspecific grapevine rootstock cultivar 'Börner' (Vitis riparia GM183 x Vitis cinerea Arnold) were used to assemble its chloroplast and mitochondrion genome sequences. We annotated 133 chloroplast and 172 mitochondrial genes including the RNA-editing sites. The organellar genomes were maternally inherited to 'Börner' from Vitis riparia. Footnotes * ORCiD added and URLs included * https://pub.uni-bielefeld.de/record/2941430 * https://pub.uni-bielefeld.de/record/2941437

The genome sequence assembly of the diploid and highly homozygous V.vinifera genotype PN40024 serves as the reference for many grapevine studies. Despite several improvements of the PN40024 genome assembly, its current version PN12X.v2 is quite fragmented and only represents the haploid state of the genome with mixed haplotypes. In fact, despite the PN40024 genome is nearly homozygous, it still contains various heterozygous regions. Taking the opportunity of the improvements that long-read sequencing technologies offer to fully discriminate haplotype sequences and considering that several Vitis sp. genomes have recently been assembled with these approaches, an improved version of the reference, called PN40024.v4, was generated. Through incorporating long genomic sequencing reads to the assembly, the continuity of the 12X.v2 scaffolds was highly increased. The number of scaffolds decreased from 2,059 to 640 and the number of N bases was reduced by 88%. Additionally, the full alternative haplotype sequence was built for the first time, the chromosome anchoring was improved and the amount of unplaced scaffolds were reduced by half. To obtain a high-quality gene annotation that outperforms previous versions, a liftover approach was complemented with an optimized annotation workflow for Vitis. Integration of the gene reference catalogue and its manual curation have also assisted in improving the annotation, while defining the most reliable estimation to date of 35,230 genes. Finally, we demonstrate that PN40024 resulted from selfings of cv. ′Helfensteiner′ (cross of cv. ′Pinot noir′ and ′Schiava grossa′) instead of a single ′Pinot noir′. These advances will help maintaining the PN40024 genome as a gold-standard reference also contributing in the eventual elaboration of the grapevine pangenome.Competing Interest StatementThe authors have declared no competing interest.