Construction of Red Fox Chromosomal Fragments from the Short-Read Genome Assembly

Abstract

The genome of a red fox (Vulpes vulpes) was recently sequenced and assembled using next-generation sequencing (NGS). The assembly is of high quality, with 94X coverage and a scaffold N50 of 11.8 Mbp, but is split into 676,878 scaffolds, some of which are likely to contain assembly errors. Fragmentation and misassembly hinder accurate gene prediction and downstream analysis such as the identification of loci under selection. Therefore, assembly of the genome into chromosome-scale fragments was an important step towards developing this genomic model. Scaffolds from the assembly were aligned to the dog reference genome and compared to the alignment of an outgroup genome (cat) against the dog to identify syntenic sequences among species. The program Reference-Assisted Chromosome Assembly (RACA) then integrated the comparative alignment with the mapping of the raw sequencing reads generated during assembly against the fox scaffolds. The 128 sequence fragments RACA assembled were compared to the fox meiotic linkage map to guide the construction of 40 chromosomal fragments. This computational approach to assembly was facilitated by prior research in comparative mammalian genomics, and the continued improvement of the red fox genome can in turn offer insight into canid and carnivore chromosome evolution. This assembly is also necessary for advancing genetic research in foxes and other canids.

Hypothalamic transcriptome of tame and aggressive silver foxes (Vulpes vulpes) identifies gene expression differences shared across brain regions

“…Finally, reads of 30 nucleotides or longer were selected and mapped to the reference genome. In the absence of a complete fox genome assembly, [36][37][38] closely related dog genome assembly (C. familiaris 3.1 genome) was used as the reference for mapping. This reference genome was also used in previous studies comparing the PFC, BFB and pituitary gland of tame vs aggressive foxes.…”

A near-chromosome-scale genome assembly of the gemsbok (Oryx gazella): an iconic antelope of the Kalahari desert

“…The other 23 breaks were inside single SOAPdenovo scaffolds, with five also being broken in the SOAPdenovo + RACA assembly, while the rest were either not used (4 cases) or below the 150 Kbp resolution of the SOAPdenovo + RACA assembly (14 cases). Although physical or genetic maps for gemsbok are not available to verify the SOAPdenovo + Chicago + RACA assembly, we previously showed that RACA produces highly accurate chromosome assemblies when compared to meiotic linkage [20] or cytogenetic physical maps [19], suggesting that the 47 PCFs of the gemsbok assembly accurately represent scaffold order and orientation on the gemsbok chromosomes. Therefore, using RACA allowed us to identify putatively chimeric scaffolds and superscaffolds, as well as to align components of chimeric scaffolds to their likely location on the gemsbok genome.…”

A Near Chromosome Assembly of the Dromedary Camel Genome

“…In this study, we upgraded the previously published fragmented dromedary camel genome assembly to nearly chromosome-level using a combination of in silico chromosome reconstructions, PCR-verification and supporting data from camel and alpaca physical maps. This approach has been previously applied to mammalian genomes, such as the Tibetan antelope (Kim et al, 2013) (2 n = 60), red fox (Rando et al, 2018) (2 n = 34), and avian species, including pigeon and peregrine falcon (Damas et al, 2017), and showed high consistency when compared with third-generation sequencing methodologies (Holt et al, 2018). Our approach resulted in a remarkable reduction in fragmentation of the original dromedary assembly by 25-fold, and an N50 increase 35-fold.…”

“…Other reference-based algorithms e.g., RAGOUT (Kolmogorov et al, 2014) do not use the target assembly read-pair data to verify scaffold structures and orders, meaning that the target species-specific rearrangements could be missed from the reconstructed chromosome fragments, which could prove to be a problem in future candidate gene research, as a lower quality genome assembly will produce more false-negative and false-positive association signals, reducing the value of association studies (Goldfeder et al, 2016). Moreover, RACA has been successfully used for other genome assembly projects, including mammals [such as Tibetan antelope and red fox (Kim et al, 2013; Rando et al, 2018)] and birds [peregrine and saker falcons, ostrich, pigeon, and budgerigar (Damas et al, 2017; O’Connor et al, 2018)]. Finally, RACA assemblies could provide an independent source to prove and/or further improve assemblies produced with such methods as HiC, 10X or Dovetail Chicago.…”