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.
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.
Breeding progress in black raspberry (Rubus occidentalis L.) has been limited by a lack of genetic diversity in elite germplasm. Black raspberry cultivars have been noted for showing very few phenotypic differences and seedlings from crosses between cultivars for a lack of segregation for important traits. Despite these challenges, little molecular work has been done to explore genetic diversity and relationships in wild and cultivated black raspberry germplasm. Microsatellite, or simple sequence repeat (SSR), markers are highly polymorphic codominant markers useful for studying genetic diversity, population genetics, genetic fingerprinting and other applications. We examined genetic diversity in 148 wild and cultivated black raspberry accessions using 21 polymorphic SSR markers. Black raspberry cultivars clustered tightly and showed higher than expected heterozygosity while that of wild accessions was low. Relationships between wild black raspberry accessions were poorly resolved and regional clusters were mostly absent from our analysis. Our results indicated that wild black raspberry germplasm is a relatively untapped resource available for future breeding.
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.
Key messageWe have constructed a densely populated, saturated genetic linkage map of black raspberry and successfully placed a locus for aphid resistance.AbstractBlack raspberry (Rubus occidentalis L.) is a high-value crop in the Pacific Northwest of North America with an international marketplace. Few genetic resources are readily available and little improvement has been achieved through breeding efforts to address production challenges involved in growing this crop. Contributing to its lack of improvement is low genetic diversity in elite cultivars and an untapped reservoir of genetic diversity from wild germplasm. In the Pacific Northwest, where most production is centered, the current standard commercial cultivar is highly susceptible to the aphid Amphorophora agathonica Hottes, which is a vector for the Raspberry mosaic virus complex. Infection with the virus complex leads to a rapid decline in plant health resulting in field replacement after only 3–4 growing seasons. Sources of aphid resistance have been identified in wild germplasm and are used to develop mapping populations to study the inheritance of these valuable traits. We have constructed a genetic linkage map using single-nucleotide polymorphism and transferable (primarily simple sequence repeat) markers for F1 population ORUS 4305 consisting of 115 progeny that segregate for aphid resistance. Our linkage map of seven linkage groups representing the seven haploid chromosomes of black raspberry consists of 274 markers on the maternal map and 292 markers on the paternal map including a morphological locus for aphid resistance. This is the first linkage map of black raspberry and will aid in developing markers for marker-assisted breeding, comparative mapping with other Rubus species, and enhancing the black raspberry genome assembly.Electronic supplementary materialThe online version of this article (doi:10.1007/s00122-015-2541-x) contains supplementary material, which is available to authorized users.
Rubus fruits are high-value crops that are sought after by consumers for their flavor, visual appeal, and health benefits. To meet this demand, production of red and black raspberries (R. idaeus L. and R. occidentalis L.), blackberries (R. subgenus Rubus), and hybrids, such as Boysenberry and marionberry, is growing worldwide. Rubus breeding programmes are continually striving to improve flavor, texture, machine harvestability, and yield, provide pest and disease resistance, improve storage and processing properties, and optimize fruits and plants for different production and harvest systems. Breeders face numerous challenges, such as polyploidy, the lack of genetic diversity in many of the elite cultivars, and until recently, the relative shortage of genetic and genomic resources available for Rubus. This review will highlight the development of continually improving genetic maps, the identification of Quantitative Trait Loci (QTL)s controlling key traits, draft genomes for red and black raspberry, and efforts to improve gene models. The development of genetic maps and markers, the molecular characterization of wild species and germplasm, and high-throughput genotyping platforms will expedite breeding of improved cultivars. Fully sequenced genomes and accurate gene models facilitate identification of genes underlying traits of interest and enable gene editing technologies such as CRISPR/Cas9.
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