These authors contributed equally to this work. SUMMARYBlack raspberry (Rubus occidentalis) is an important specialty fruit crop in the US Pacific Northwest that can hybridize with the globally commercialized red raspberry (R. idaeus). Here we report a 243 Mb draft genome of black raspberry that will serve as a useful reference for the Rosaceae and Rubus fruit crops (raspberry, blackberry, and their hybrids). The black raspberry genome is largely collinear to the diploid woodland strawberry (Fragaria vesca) with a conserved karyotype and few notable structural rearrangements. Centromeric satellite repeats are widely dispersed across the black raspberry genome, in contrast to the tight association with the centromere observed in most plants. Among the 28 005 predicted protein-coding genes, we identified 290 very recent small-scale gene duplicates enriched for sugar metabolism, fruit development, and anthocyanin related genes which may be related to key agronomic traits during black raspberry domestication. This contrasts patterns of recent duplications in the wild woodland strawberry F. vesca, which show no patterns of enrichment, suggesting gene duplications contributed to domestication traits. Expression profiles from a fruit ripening series and roots exposed to Verticillium dahliae shed insight into fruit development and disease response, respectively. The resources presented here will expedite the development of improved black and red raspberry, blackberry and other Rubus cultivars.

the importance of using multiple phylogenetic methods when examining complex groups and the utility of software programs that estimate signal conflict within datasets.

BackgroundComparative genome mapping studies in Rosaceae have been conducted until now by aligning genetic maps within the same genus, or closely related genera and using a limited number of common markers. The growing body of genomics resources and sequence data for both Prunus and Fragaria permits detailed comparisons between these genera and the recently released Malus × domestica genome sequence.ResultsWe generated a comparative analysis using 806 molecular markers that are anchored genetically to the Prunus and/or Fragaria reference maps, and physically to the Malus genome sequence. Markers in common for Malus and Prunus, and Malus and Fragaria, respectively were 784 and 148. The correspondence between marker positions was high and conserved syntenic blocks were identified among the three genera in the Rosaceae. We reconstructed a proposed ancestral genome for the Rosaceae.ConclusionsA genome containing nine chromosomes is the most likely candidate for the ancestral Rosaceae progenitor. The number of chromosomal translocations observed between the three genera investigated was low. However, the number of inversions identified among Malus and Prunus was much higher than any reported genome comparisons in plants, suggesting that small inversions have played an important role in the evolution of these two genera or of the Rosaceae.

BackgroundThe fragmented nature of most draft plant genomes has hindered downstream gene discovery, trait mapping for breeding, and other functional genomics applications. There is a pressing need to improve or finish draft plant genome assemblies.FindingsHere, we present a chromosome-scale assembly of the black raspberry genome using single-molecule real-time Pacific Biosciences sequencing and high-throughput chromatin conformation capture (Hi-C) genome scaffolding. The updated V3 assembly has a contig N50 of 5.1 Mb, representing an ∼200-fold improvement over the previous Illumina-based version. Each of the 235 contigs was anchored and oriented into seven chromosomes, correcting several major misassemblies. Black raspberry V3 contains 47 Mb of new sequences including large pericentromeric regions and thousands of previously unannotated protein-coding genes. Among the new genes are hundreds of expanded tandem gene arrays that were collapsed in the Illumina-based assembly. Detailed comparative genomics with the high-quality V4 woodland strawberry genome (Fragaria vesca) revealed near-perfect 1:1 synteny with dramatic divergence in tandem gene array composition. Lineage-specific tandem gene arrays in black raspberry are related to agronomic traits such as disease resistance and secondary metabolite biosynthesis.ConclusionsThe improved resolution of tandem gene arrays highlights the need to reassemble these highly complex and biologically important regions in draft plant genomes. The updated, high-quality black raspberry reference genome will be useful for comparative genomics across the horticulturally important Rosaceae family and enable the development of marker assisted breeding in Rubus.

A linkage mapping approach was used to identify quantitative trait loci (QTL) associated with day-neutrality in the commercial strawberry, Fragaria · ananassa (Duch ex Rozier). Amplified Fragment Length Polymorphism (AFLP) markers were used to build a genetic map with a population of 127 lines developed by crossing the dayneutral (DN) ÔTributeÕ with the short-day (SD) ÔHoneoyeÕ. The population was genotyped with AFLP markers and 429 single dose restriction fragments (SDRF) were placed on a consensus map of 1541 cM with 43 linkage groups. Individuals from the mapping population were observed for their flowering habit throughout the growing season in Michigan (MI), Minnesota (MN), Maryland (MD), Oregon (OR) and California (CA). Eight QTL were found that were either location specific or shared among locations. None of these QTL explained >36% of the phenotypic variation, indicating that the inheritance of day-neutrality is likely a polygenic trait.Two primary types of commercial strawberries are grown, short-day (SD) and day-neutral (DN). SD genotypes or Junebearers, initiate flower buds either under SD conditions (<14 h of day length) or at temperatures below 15°C, while DN genotypes are photoperiod insensitive and will initiate flowers under any photoperiod conditions as long as temperatures are moderate (Darrow 1966, Hancock 1999. Dayneutrality was most recently introduced into modern cultivars by Bringhurst and Voth (1984), using a native genotype of F. virginiana (Mill) ssp. glauca (S. Watson) Staudt from the Wasatch Mountains of Utah.To date, the genetics of day-neutrality in strawberries have remained elusive. Several different models have been proposed including: (i) regulation by a single dominant gene Voth 1978, Ahmadi et al. 1990); (ii) regulation by dominant complementary genes (Ourecky and Slate 1967); and (iii) quantitative inheritance (Powers 1954, Hancock et al. 2001). The reason why these studies generated different hypotheses may be that they utilized different sets of parents and were conducted in different environments. The study of Ourecky and Slate (1967) was conducted in New York using material that had not recently had any new F. virginiana germplasm incorporated. The studies of Powers (1954) and Hancock et al. (2001), were performed in Wyoming and Michigan, respectively, using DN parents that carried genes from F. · ananassa and wild clones of F. virginiana that were different from the Wasatch source. The studies of Bringhurst and Voth (1978) and Ahmadi et al. (1990) were performed in CA using University of California-Davis breeding parents carrying the Wasatch source of day-neutrality. There was one study in CA that suggested day-neutrality may have a quantitative basis (Shaw 2003), but it was later refuted by a more extensive statistical analysis of a greater number of progeny populations (Shaw and Famula 2005). Sugimoto et al. (2005) found a RAPD-marker linked to a dominant gene regulating day-neutrality in a Japanese breeding population carrying the Wasatch source of day-neutrality.To e...

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Black raspberry (Rubus occidentalis L.) is a niche fruit crop valued for its flavor and potential health benefits. The improvement of fruit and cane characteristics via molecular breeding technologies has been hindered by the lack of a high-quality reference genome. The recently released draft genome for black raspberry (ORUS 4115-3) lacks assembly of scaffolds to chromosome scale. We used high-throughput chromatin conformation capture (Hi-C) and Proximity-Guided Assembly (PGA) to cluster and order 9650 out of 11,936 contigs of this draft genome assembly into seven pseudo-chromosomes. The seven pseudo-chromosomes cover ~97.2% of the total contig length (~223.8 Mb). Locating existing genetic markers on the physical map resolved multiple discrepancies in marker order on the genetic map. Centromeric regions were inferred from recombination frequencies of genetic markers, alignment of 303 bp centromeric sequence with the PGA, and heat map showing the physical contact matrix over the entire genome. We demonstrate a high degree of synteny between each of the seven chromosomes of black raspberry and a high-quality reference genome for strawberry (Fragaria vesca L.) assembled using only PacBio long-read sequences. We conclude that PGA is a cost-effective and rapid method of generating chromosome-scale assemblies from Illumina short-read sequencing data.

The genus Rubus belongs to the Rosaceae and is comprised of 600-800 species distributed world-wide. To date, genetic maps of the genus consist largely of non-transferable markers such as amplified fragment length polymorphisms. An F(1) population developed from a cross between an advanced breeding selection of Rubus occidentalis (96395S1) and R. idaeus 'Latham' was used to construct a new genetic map consisting of DNA sequence-based markers. The genetic linkage maps presented here are constructed of 131 markers on at least one of the two parental maps. The majority of the markers are orthologous, including 14 Rosaceae conserved orthologous set markers, and 60 new gene-based markers developed for raspberry. Thirty-four published raspberry simple sequence repeat markers were used to align the new maps to published raspberry maps. The 96395S1 genetic map consists of six linkage groups (LG) and covers 309 cM with an average of 10 cM between markers; the 'Latham' genetic map consists of seven LG and covers 561 cM with an average of 5 cM between markers. We used BLAST analysis to align the orthologous sequences used to design primer pairs for Rubus genetic mapping with the genome sequences of Fragaria vesca 'Hawaii 4', Malus × domestica 'Golden Delicious', and Prunus 'Lovell'. The alignment of the orthologous markers designed here suggests that the genomes of Rubus and Fragaria have a high degree of synteny and that synteny decreases with phylogenetic distance. Our results give unprecedented insights into the genome evolution of raspberry from the putative ancestral genome of the single ancestor common to Rosaceae.