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Dog domestication involved long-term genetic selection for behavior. The genetics-centered view of domestication is supported by experimental selection of farm-foxes (Vulpes vulpes) that began in the 1950s. Selection of foxes, separately, for tame and for aggressive behavior, has yielded two strains with markedly different, genetically determined behavioral phenotypes. Tame-strain foxes communicate with humans in a positive manner and are eager to establish human contact. Foxes from aggressive strains are aggressive to humans and difficult to handle. Although selected solely for behavior, changes in physiology, morphology, and appearance with significant parallels to characteristics of the domestic dog, were observed in tame-strain. Ongoing research is focused on identification of molecular genetic mechanisms associated with selection of foxes for behavior. Identification of behavioral loci in the fox genome in the region which is homologous to the region in the dog genome that differentiates dogs from wolves lead to the hypothesis that domesticated behavior in dogs and foxes may have similar genetic bases.

The red fox (Vulpes vulpes) has a wide global distribution with many ecotypes, and has been bred in captivity for various traits, making it a useful evolutionary model system. The Y chromosome represents one of the most informative markers of phylogeography, yet it has not been well-studied in the red fox due to a lack of the necessary genomic resources. We used a target capture approach to sequence a portion of the red fox Y chromosome in a geographically diverse red fox sample, along with other canid species, to develop single nucleotide polymorphism (SNP) markers, 13 of which we validated for use in subsequent studies. Phylogenetic analyses of the Y chromosome sequences, including calibration to outgroups, confirmed previous estimates of the timing of two intercontinental exchanges of red foxes, the initial colonization of North America from Eurasia approximately half a million years ago and a subsequent continental exchange before the last Pleistocene glaciation (∼100,000 years ago). However, in contrast to mtDNA, which showed unidirectional transfer from Eurasia to North America prior to the last glaciation, the Y chromosome appears to have been transferred from North America to Eurasia during this period. Additional sampling is needed to confirm this pattern and to further clarify red fox Y chromosome phylogeography.

Strains of red fox (Vulpes vulpes) with markedly different behavioural phenotypes have been developed in the famous long-term selective breeding programme known as the Russian farm-fox experiment. Here we sequenced and assembled the red fox genome and re-sequenced a subset of foxes from the tame, aggressive and conventional farm-bred populations to identify genomic regions associated with the response to selection for behaviour. Analysis of the re-sequenced genomes identified 103 regions with either significantly decreased heterozygosity in one of the three populations or increased divergence between the populations. A strong positional candidate gene for tame behaviour was highlighted: SorCS1, which encodes the main trafficking protein for AMPA glutamate receptors and neurexins and suggests a role for synaptic plasticity in fox domestication. Other regions identified as likely to have been under selection in foxes include genes implicated in human neurological disorders, mouse behaviour and dog domestication. The fox represents a powerful model for the genetic analysis of affiliative and aggressive behaviours that can benefit genetic studies of behaviour in dogs and other mammals, including humans.

Domesticated species exhibit a suite of behavioral, endocrinological, and morphological changes referred to as “domestication syndrome.” These changes may include a reduction in reactivity of the hypothalamic-pituitary-adrenal (HPA) axis and specifically reduced adrenocorticotropic hormone release from the anterior pituitary. To investigate the biological mechanisms targeted during domestication, we investigated gene expression in the pituitaries of experimentally domesticated foxes (Vulpes vulpes). RNA was sequenced from the anterior pituitary of six foxes selectively bred for tameness (“tame foxes”) and six foxes selectively bred for aggression (“aggressive foxes”). Expression, splicing, and network differences identified between the two lines indicated the importance of genes related to regulation of exocytosis, specifically mediated by cAMP, organization of pseudopodia, and cell motility. These findings provide new insights into biological mechanisms that may have been targeted when these lines of foxes were selected for behavior and suggest new directions for research into HPA axis regulation and the biological underpinnings of domestication.

Pathogens are recognized as major drivers of local adaptation in wildlife systems. By determining which gene variants are favored in local interactions among populations with and without disease, spatially explicit adaptive responses to pathogens can be elucidated. Much of our current understanding of host responses to disease comes from a small number of genes associated with an immune response. High‐throughput sequencing (HTS) technologies, such as genotype‐by‐sequencing (GBS), facilitate expanded explorations of genomic variation among populations. Hybridization‐based GBS techniques can be leveraged in systems not well characterized for specific variants associated with disease outcome to “capture” specific genes and regulatory regions known to influence expression and disease outcome. We developed a multiplexed, sequence capture assay for red foxes to simultaneously assess ~300‐kbp of genomic sequence from 116 adaptive, intrinsic, and innate immunity genes of predicted adaptive significance and their putative upstream regulatory regions along with 23 neutral microsatellite regions to control for demographic effects. The assay was applied to 45 fox DNA samples from Alaska, where three arctic rabies strains are geographically restricted and endemic to coastal tundra regions, yet absent from the boreal interior. The assay provided 61.5% on‐target enrichment with relatively even sequence coverage across all targeted loci and samples (mean = 50×), which allowed us to elucidate genetic variation across introns, exons, and potential regulatory regions (4,819 SNPs). Challenges remained in accurately describing microsatellite variation using this technique; however, longer‐read HTS technologies should overcome these issues. We used these data to conduct preliminary analyses and detected genetic structure in a subset of red fox immune‐related genes between regions with and without endemic arctic rabies. This assay provides a template to assess immunogenetic variation in wildlife disease systems.

The underlying neurological events accompanying dog domestication remain elusive. To reconstruct the domestication process in an experimental setting, silver foxes (Vulpes vulpes) have been deliberately bred for tame vs aggressive behaviors for more than 50 generations at the Institute for Cytology and Genetics in Novosibirsk, Russia. The hypothalamus is an essential part of the hypothalamic-pituitary-adrenal axis and regulates the fight-or-flight response, and thus, we hypothesized that selective breeding for tameness/aggressiveness has shaped the hypothalamic transcriptomic profile. RNA-seq analysis identified 70 differentially expressed genes (DEGs). Seven of these genes, DKKL1, FBLN7, NPL, PRIMPOL, PTGRN, SHCBP1L and SKIV2L, showed the same direction expression differences in the hypothalamus, basal forebrain and prefrontal cortex. The genes differentially expressed across the three tissues are involved in cell division, differentiation, adhesion and carbohydrate processing, suggesting an association of these processes with selective breeding. Additionally, 159 transcripts from the hypothalamus demonstrated differences in the abundance of alternative spliced forms between the tame and aggressive foxes. Weighted gene coexpression network analyses also suggested that gene modules in hypothalamus were significantly associated with tame vs aggressive behavior. Pathways associated with these modules include signal transduction, interleukin signaling, cytokine-cytokine receptor interaction and peptide ligand-binding receptors (eg, G-protein coupled receptor [GPCR] ligand binding). Current studies show the selection for tameness vs aggressiveness in foxes is associated with unique hypothalamic gene profiles partly shared with other brain regions and highlight DEGs involved in biological processes such as development, differentiation and immunological responses. The role of these processes in fox and dog domestication remains to be determined.

While the number of mammalian genome assemblies has proliferated, Y-chromosome assemblies have lagged behind. This discrepancy is caused by biological features of the Y-chromosome, such as its high repeat content, that present challenges to assembly with short-read, next-generation sequencing technologies. Partial Y-chromosome assemblies have been developed for the cat (Felis catus), dog (Canis lupus familiaris), and grey wolf (Canis lupus lupus), providing the opportunity to examine the red fox (Vulpes vulpes) Y-chromosome in the context of closely related species. Here we present a data-driven approach to identifying Y-chromosome sequence among the scaffolds that comprise the short-read assembled red fox genome. First, scaffolds containing genes found on the Y-chromosomes of cats, dogs, and wolves were identified. Next, analysis of the resequenced genomes of 15 male and 15 female foxes revealed scaffolds containing male-specific k-mers and patterns of inter-sex copy number variation consistent with the heterogametic chromosome. Analyzing variation across these two metrics revealed 171 scaffolds containing 3.37 Mbp of putative Y-chromosome sequence. The gene content of these scaffolds is consistent overall with that of the Y-chromosome in other carnivore species, though the red fox Y-chromosome carries more copies of BCORY2 and UBE1Y than has been reported in related species and fewer copies of SRY than in other canids. The assignment of these scaffolds to the Y-chromosome serves to further characterize the content of the red fox draft genome while providing resources for future analyses of canid Y-chromosome evolution.

The role of chromosome rearrangements in driving evolution has been a long-standing question of evolutionary biology. Here we focused on ruminants as a model to assess how rearrangements may have contributed to the evolution of gene regulation. Using reconstructed ancestral karyotypes of Cetartiodactyls, Ruminants, Pecorans, and Bovids, we traced patterns of gross chromosome changes. We found that the lineage leading to the ruminant ancestor after the split from other cetartiodactyls was characterized by mostly intrachromosomal changes, whereas the lineage leading to the pecoran ancestor (including all livestock ruminants) included multiple interchromosomal changes. We observed that the liver cell putative enhancers in the ruminant evolutionary breakpoint regions are highly enriched for DNA sequences under selective constraint acting on lineage-specific transposable elements (TEs) and a set of 25 specific transcription factor (TF) binding motifs associated with recently active TEs. Coupled with gene expression data, we found that genes near ruminant breakpoint regions exhibit more divergent expression profiles among species, particularly in cattle, which is consistent with the phylogenetic origin of these breakpoint regions. This divergence was significantly greater in genes with enhancers that contain at least one of the 25 specific TF binding motifs and located near bovidae-to-cattle lineage breakpoint regions. Taken together, by combining ancestral karyotype reconstructions with analysis of cis regulatory element and gene expression evolution, our work demonstrated that lineage-specific regulatory elements colocalized with gross chromosome rearrangements may have provided valuable functional modifications that helped to shape ruminant evolution.

The Russian farm-fox experiment is an unusually long-running and well-controlled study designed to replicate wolf-to-dog domestication. As such, it offers an unprecedented window onto the neural mechanisms governing the evolution of behavior. Here we report evolved changes to gray matter morphology resulting from selection for tameness versus aggressive responses toward humans in a sample of 30 male fox brains. Contrasting with standing ideas on the effects of domestication on brain size, tame foxes did not show reduced brain volume. Rather, gray matter volume in both the tame and aggressive strains was increased relative to conventional farm foxes bred without deliberate selection on behavior. Furthermore, tame- and aggressive-enlarged regions overlapped substantially, including portions of motor, somatosensory, and prefrontal cortex, amygdala, hippocampus, and cerebellum. We also observed differential morphologic covariation across distributed gray matter networks. In one prefrontal-cerebellum network, this covariation differentiated the three populations along the tame-aggressive behavioral axis. Surprisingly, a prefrontal-hypothalamic network differentiated the tame and aggressive foxes together from the conventional strain. These findings indicate that selection for opposite behaviors can influence brain morphology in a similar way. SIGNIFICANCE STATEMENT Domestication represents one of the largest and most rapid evolutionary shifts of life on earth. However, its neural correlates are largely unknown. Here we report the neuroanatomical consequences of selective breeding for tameness or aggression in the seminal Russian farm-fox experiment. Compared with a population of conventional farm-bred control foxes, tame foxes show neuroanatomical changes in the PFC and hypothalamus, paralleling wolf-to-dog shifts. Surprisingly, though, aggressive foxes also show similar changes. Moreover, both strains show increased gray matter volume relative to controls. These results indicate that similar brain adaptations can result from selection for opposite behavior, that existing ideas of brain changes in domestication may need revision, and that significant neuroanatomical change can evolve very quickly, within the span of

Individuals involved in a social interaction exhibit different behavioral traits that, in combination, form the individual’s behavioral responses. Selectively bred strains of silver foxes (Vulpes vulpes) demonstrate markedly different behaviors in their response to humans. To identify the genetic basis of these behavioral differences we constructed a large F2 population including 537 individuals by cross-breeding tame and aggressive fox strains. 98 fox behavioral traits were recorded during social interaction with a human experimenter in a standard four-step test. Patterns of fox behaviors during the test were evaluated using principal component (PC) analysis. Genetic mapping identified eight unique significant and suggestive QTL. Mapping results for the PC phenotypes from different test steps showed little overlap suggesting that different QTL are involved in regulation of behaviors exhibited in different behavioral contexts. Many individual behavioral traits mapped to the same genomic regions as PC phenotypes. This provides additional information about specific behaviors regulated by these loci. Further, three pairs of epistatic loci were also identified for PC phenotypes suggesting more complex genetic architecture of the behavioral differences between the two strains than what has previously been observed.

Animal domestication efforts have led to a shared spectrum of striking behavioral and morphological changes. To recapitulate this process, silver foxes have been selectively bred for tame and aggressive behaviors for more than 50 generations at the Institute for Cytology and Genetics in Novosibirsk, Russia. To understand the genetic basis and molecular mechanisms underlying the phenotypic changes, we profiled gene expression levels and coding SNP allele frequencies in two brain tissue specimens from 12 aggressive foxes and 12 tame foxes. Expression analysis revealed 146 genes in the prefrontal cortex and 33 genes in the basal forebrain that were differentially expressed, with a 5% false discovery rate (FDR). These candidates include genes in key pathways known to be critical to neurologic processing, including the serotonin and glutamate receptor pathways. In addition, 295 of the 31,000 exonic SNPs show significant allele frequency differences between the tame and aggressive populations (1% FDR), including genes with a role in neural crest cell fate determination.

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.

The de novo assembly of the red fox (Vulpes vulpes) genome has facilitated the development of genomic tools for the species. Efforts to identify the population history of red foxes in North America have previously been limited by a lack of information about the red fox Y-chromosome sequence. However, a megabase of red fox Y-chromosome sequence was recently identified over 2 scaffolds in the reference genome. Here, these scaffolds were scanned for repeated motifs, revealing 194 likely microsatellites. Twenty-three of these loci were selected for primer development and, after testing, produced a panel of 11 novel markers that were analyzed alongside 2 markers previously developed for the red fox from dog Y-chromosome sequence. The markers were genotyped in 76 male red foxes from 4 populations: 7 foxes from Newfoundland (eastern Canada), 12 from Maryland (eastern United States), and 9 from the island of Great Britain, as well as 48 foxes of known North American origin maintained on an experimental farm in Novosibirsk, Russia. The full marker panel revealed 22 haplotypes among these red foxes, whereas the 2 previously known markers alone would have identified only 10 haplotypes. The haplotypes from the 4 populations clustered primarily by continent, but unidirectional gene flow from Great Britain and farm populations may influence haplotype diversity in the Maryland population. The development of new markers has increased the resolution at which red fox Y-chromosome diversity can be analyzed and provides insight into the contribution of males to red fox population diversity and patterns of phylogeography.

Although animal domestication has enjoyed the attention of geneticists and evolutionary biologists since Darwin’s times, the focal question about the genetic basis of this process has not been duly considered. We present some results of experimental modeling of historical domestication in silver foxes (Vulpes vulpes), an object of commercial breeding. Attention is focused on the role of artificial selection in the transformation of the behavior of these animals, which has brought about unique populations of tame and aggressive foxes. Additional resources were developed to analyze the molecular nature of differences in the behavior of these unique foxes: the meiotic map of the fox genome was constructed, informative segregating subpopulations were obtained by crosses of tame and aggressive animals, and the formerly applied method of behavioral phenotype quantification was improved. The integrated behavioral phenotypes (principal components PC1 and PC2) used in the study were obtained by analysis of ethological parameters recorded with a camcorder. The most important result is that the region most closely associated with tame behavior was identified on fox chromosome 12 by QTL interval mapping. The principle significance of this result is strengthened by the fact that the region is similar to the region on dog chromosome 5, presumed to be responsible for the early domestication of wolves and their evolutionary transformation to primitive dogs.

The foxes at Novosibirsk, Russia, are the only population of domesticated foxes in the world. These domesticated foxes originated from farm-bred silver foxes (Vulpes vulpes), whose genetic source is unknown. In this study we examined the origin of the domesticated strain of foxes and two other farm-bred fox populations (aggressive and unselected) maintained in Novosibirsk. To identify the phylogenetic origin of these populations we sequenced two regions of mtDNA, cytochrome b and D-loop, from 24 Novosibirsk foxes (8 foxes from each population) and compared them with corresponding sequences of native red foxes from Europe, Asia, Alaska and Western Canada, Eastern Canada, and the Western Mountains of the USA. We identified seven cytochrome b - D-loop haplotypes in Novosibirsk populations, four of which were previously observed in Eastern North America. The three remaining haplotypes differed by one or two base change from the most common haplotype in Eastern Canada. Φ(ST) analysis showed significant differentiation between Novosibirsk populations and red fox populations from all geographic regions except Eastern Canada. No haplotypes of Eurasian origin were identified in the Novosibirsk populations. These results are consistent with historical records indicating that the original breeding stock of farm-bred foxes originated from Prince Edward Island, Canada. Mitochondrial DNA data together with historical records indicate two stages in the selection of domesticated foxes: the first includes captive breeding for ~50 years with unconscious selection for behaviour; the second corresponds to over 50 further years of intensive selection for tame behaviour.

Fine mapping followed by candidate gene analysis of erd — a canine hereditary retinal degeneration characterized by aberrant photoreceptor development — established that the disease cosegregates with a SINE insertion in exon 4 of the canine STK38L/NDR2 gene. The mutation removes exon 4 from STK38L transcripts and is predicted to remove much of the N terminus from the translated protein, including binding sites for S100B and Mob proteins, part of the protein kinase domain, and a Thr-75 residue critical for autophosphorylation. Although known to have roles in neuronal cell function, the STK38L pathway has not previously been implicated in normal or abnormal photoreceptor development. Loss of STK38L function in erd provides novel potential insights into the role of the STK38L pathway in neuronal and photoreceptor cell function, and suggests that genes in this pathway need to be considered as candidate genes for hereditary retinal degenerations.

In the version of this Article originally published, there were some errors in the affiliations: Stephen J. O’Brien’s affiliations were incorrectly listed as 8,9; they should have been 7,9. Affiliation 3 was incorrectly named the Institute of Cytology and Genetics of the Russian Academy of Sciences; it should have read Institute of Cytology and Genetics of the Siberian Branch of the Russian Academy of Sciences. Affiliation 4 was incorrectly named the Institute of Molecular and Cell Biology of the Russian Academy of Sciences; it should have read Institute of Molecular and Cellular Biology of the Siberian Branch of the Russian Academy of Sciences. These have now been corrected.

B chromosomes (Bs) represent a variable addition to the main karyotype in some lineages of animals and plants. Bs accumulate through non-Mendelian inheritance and become widespread in populations. Despite the presence of multiple genes, most Bs lack specific phenotypic effects, although their influence on host genome epigenetic status and gene expression are recorded. Previously, using sequencing of isolated Bs of ruminants and rodents, we demonstrated that Bs originate as segmental duplications of specific genomic regions, and subsequently experience pseudogenization and repeat accumulation. Here, we used a similar approach to characterize Bs of the red fox (Vulpes vulpes L.) and the Chinese raccoon dog (Nyctereutes procyonoides procyonoides Gray). We confirm the previous findings of the KIT gene on Bs of both species, but demostrate an independent origin of Bs in these species, with two reused regions. Comparison of gene ensembles in Bs of canids, ruminants, and rodents once again indicates enrichment with cell-cycle genes, development-related genes, and genes functioning in the neuron synapse. The presence of B-chromosomal copies of genes involved in cell-cycle regulation and tissue differentiation may indicate importance of these genes for B chromosome establishment.

Oculo-skeletal dysplasia segregates in two canine breeds, the Labrador retriever and samoyed, in which the causative loci have been termed drd1 and drd2, respectively. Affected dogs exhibit short-limbed dwarfism together with severe ocular defects, and this phenotype is inherited as an autosomal recessive trait in both breeds. The clinical and pathological appearance resembles human hereditary arthro-ophthalmopathies such as Stickler syndrome, or Marshall Syndrome, although these human disorders are usually dominant. Linkage studies in drd1-informative pedigrees mapped the locus to canine chromosome 24, and led to the identification of an insertional mutation in exon 1 of the gene COL9A3 that cosegregates with the disease. The drd2 locus was similarly mapped to canine chromosome 15 and shown to cosegregate with a 1,267 bp deletion mutation in the 5′ end of COL9A2. Both mutations affect the COL3 domain of the respective gene. Northern analysis showed reduced RNA expression in affected retina compared to normal. These models offer potential for studies such as protein-protein interactions between different members of the collagen gene family; regulation and expression of these genes in retina and cartilage, and even opportunities for gene therapy.

B chromosomes are often considered to be one of the most mysterious elements of karyotypes (Camacho, 2004). It is generally believed that mammalian B chromosomes do not contain any protein coding genes. The discovery of a conserved KIT gene in Canidae B chromosomes has changed this view. Here we performed analysis of sequences surrounding KIT in B chromosomes of the fox and raccoon dog. The presence of the RPL23A pseudogene was shown in canid B chromosomes. The 3′ end fragment of the KDR gene was found in raccoon dog B chromosomes. The size of the B-specific fragment homologous to the autosome fragment was estimated to be a minimum of 480 kbp in both species. The origin and evolution of B chromosomes in Canidae are discussed.

The silver fox (Vulpes vulpes) offers a novel model for studying the genetics of social behavior and animal domestication. Selection of foxes, separately, for tame and for aggressive behavior has yielded two strains with markedly different, genetically determined, behavioral phenotypes. Tame strain foxes are eager to establish human contact while foxes from the aggressive strain are aggressive and difficult to handle. These strains have been maintained as separate outbred lines for over 40 generations but their genetic structure has not been previously investigated. We applied a genotyping-by-sequencing (GBS) approach to provide insights into the genetic composition of these fox populations. Sequence analysis of EcoT22I genomic libraries of tame and aggressive foxes identified 48,294 high quality SNPs. Population structure analysis revealed genetic divergence between the two strains and more diversity in the aggressive strain than in the tame one. Significant differences in allele frequency between the strains were identified for 68 SNPs. Three of these SNPs were located on fox chromosome 14 within an interval of a previously identified behavioral QTL, further supporting the importance of this region for behavior. The GBS SNP data confirmed that significant genetic diversity has been preserved in both fox populations despite many years of selective breeding. Analysis of SNP allele frequencies in the two populations identified several regions of genetic divergence between the tame and aggressive foxes, some of which may represent targets of selection for behavior. The GBS protocol used in this study significantly expanded genomic resources for the fox, and can be adapted for SNP discovery and genotyping in other canid species.

Canine early retinal degeneration (erd) is an early onset form of canine progressive retinal atrophy phenotypically similar to human retinitis pigmentosa. In a previous study, the locus responsible for erd was mapped to canine chromosome 27 in the region corresponding to HSA12p, a region where no human retinal degeneration loci have been mapped. Canine SHARP1 gene has been localized on CFA27 in the erd interval by RH mapping, and considered as a positional candidate gene for erd. SHARP1 was cloned and sequenced from normal and erd affected dogs, and no disease-causing mutations were identified. Genotyping of 117 dogs from informative pedigrees did not reveal any recombinants between SHARP1 and erd. To date SHARP1 gene is the closest gene-specific marker to erd; genotyping additional informative pedigrees, and sequencing SHARP1 upstream regions from normal and affected dogs will be necessary to establish if SHARP1 is involved in this canine retinal disease.

Collie eye anomaly (cea) is a hereditary ocular disorder affecting development of the choroid and sclera segregating in several breeds of dog, including rough, smooth, and Border collies and Australian shepherds. The disease is reminiscent of the choroidal hypoplasia phenotype observed in humans in conjunction with craniofacial or renal abnormalities. In dogs, however, the clinical phenotype can vary significantly; many dogs exhibit no obvious clinical consequences and retain apparently normal vision throughout life, while severely affected animals develop secondary retinal detachment, intraocular hemorrhage, and blindness. We report genetic studies establishing that the primary cea phenotype, choroidal hypoplasia, segregates as an autosomal recessive trait with nearly 100% penetrance. We further report linkage mapping of the primary cea locus to a 3.9-cM region of canine chromosome 37 (LOD = 22.17 at theta = 0.076), in a region corresponding to human chromosome 2q35. These results suggest the presence of a developmental regulatory gene important in ocular embryogenesis, with potential implications for other disorders of ocular vascularization.

The phenomenon of a remarkable conservation of the X chromosome in eutherian mammals has been first described by Susumu Ohno in 1964. A notable exception is the cetartiodactyl X chromosome, which varies widely in morphology and G-banding pattern between species. It is hypothesized that this sex chromosome has undergone multiple rearrangements that changed the centromere position and the order of syntenic segments over the last 80 million years of Cetartiodactyla speciation. To investigate its evolution we have selected 26 evolutionarily conserved bacterial artificial chromosome (BAC) clones from the cattle CHORI-240 library evenly distributed along the cattle X chromosome. High-resolution BAC maps of the X chromosome on a representative range of cetartiodactyl species from different branches: pig (Suidae), alpaca (Camelidae), gray whale (Cetacea), hippopotamus (Hippopotamidae), Java mouse-deer (Tragulidae), pronghorn (Antilocapridae), Siberian musk deer (Moschidae), and giraffe (Giraffidae) were obtained by fluorescent in situ hybridization. To trace the X chromosome evolution during fast radiation in specious families, we performed mapping in several cervids (moose, Siberian roe deer, fallow deer, and Pere David's deer) and bovid (muskox, goat, sheep, sable antelope, and cattle) species. We have identified three major conserved synteny blocks and rearrangements in different cetartiodactyl lineages and found that the recently described phenomenon of the evolutionary new centromere emergence has taken place in the X chromosome evolution of Cetartiodactyla at least five times. We propose the structure of the putative ancestral cetartiodactyl X chromosome by reconstructing the order of syntenic segments and centromere position for key groups.

The red fox (Vulpes vulpes) demonstrates a variety of coat colors including platinum, a common phenotype maintained in farm-bred fox populations. Foxes heterozygous for the platinum allele have a light silver coat and extensive white spotting, whereas homozygosity is embryonic lethal. Two KIT transcripts were identified in skin cDNA from platinum foxes. The long transcript was identical to the KIT transcript of silver foxes, whereas the short transcript, which lacks exon 17, was specific to platinum. The KIT gene has several copies in the fox genome: an autosomal copy on chromosome 2 and additional copies on the B chromosomes. To identify the platinum-specific KIT sequence, the genomes of one platinum and one silver fox were sequenced. A single nucleotide polymorphism (SNP) was identified at the first nucleotide of KIT intron 17 in the platinum fox. In platinum foxes, the A allele of the SNP disrupts the donor splice site and causes exon 17, which is part of a segment that encodes a conserved tyrosine kinase domain, to be skipped. Complete cosegregation of the A allele with the platinum phenotype was confirmed by linkage mapping (LOD 25.59). All genotyped farm-bred platinum foxes from Russia and the US were heterozygous for the SNP (A/G), whereas foxes with different coat colors were homozygous for the G allele. Identification of the platinum mutation suggests that other fox white-spotting phenotypes, which are allelic to platinum, would also be caused by mutations in the KIT gene.

Background There is a growing body of evidence that B chromosomes, once regarded as totally heterochromatic and genetically inert, harbor multiple segmental duplications containing clusters of ribosomal RNA genes, processed pseudogenes and protein-coding genes. Application of novel molecular approaches further supports complex composition and possible phenotypic effects of B chromosomes. Results Here we review recent findings of gene-carrying genomic segments on B chromosomes from different vertebrate groups. We demonstrate that the genetic content of B chromosomes is highly heterogeneous and some B chromosomes contain multiple large duplications derived from various chromosomes of the standard karyotype. Although B chromosomes seem to be mostly homologous to each other within a species, their genetic content differs between species. There are indications that some genomic regions are more likely to be located on B chromosomes. Conclusions The discovery of multiple autosomal genes on B chromosomes opens a new discussion about their possible effects ranging from sex determination to fitness and adaptation, their complex interactions with host genome and role in evolution.

The ftsZ gene was found among representatives of all bacterial groups. FtsZ protein is an essential component of cell division ring. Contraction of this cytoskeleton-like ring is believed to be the universal way of bacterial division. Acholeplasma laidlawii possesses all features of the minimal mycoplasma cell and some traits of cell-wall bacteria and seems to be a promising object for study of basic principles of the bacterial division process. We cloned an A. laidlawii chromosomal fragment containing ftsZ gene and two flanking orf which also were identified. A. laidlawii FtsZ protein has been determined with polyclonal antibodies raised in rabbit. It was demonstrated that ftsZ gene of A. laidlawii could be expressed in E. coli cells. We also revealed that A. laidlawii FtsZ had a low similarity to proteins of Mycoplasma genitalium and M. pneumoniae. The comparison of FtsZ structures may be used for investigation of bacterial phylogenetic relations.

Background Achromatopsia is an autosomal recessive disease characterized by the loss of cone photoreceptor function that results in day-blindness, total colorblindness, and decreased central visual acuity. The most common causes for the disease are mutations in the CNGB3 gene, coding for the beta subunit of the cyclic nucleotide-gated channels in cones. CNGB3-achromatopsia, or cone degeneration (cd), is also known to occur in two canine breeds, the Alaskan malamute (AM) and the German shorthaired pointer. Results Here we report an in-depth characterization of the achromatopsia phenotype in a new canine breed, the miniature Australian shepherd (MAS). Genotyping revealed that the dog was homozygous for a complete genomic deletion of the CNGB3 gene, as has been previously observed in the AM. Identical breakpoints on chromosome 29 were identified in both the affected AM and MAS with a resulting deletion of 404,820 bp. Pooled DNA samples of unrelated purebred Australian shepherd, MAS, Siberian husky, Samoyed and Alaskan sled dogs were screened for the presence of the affected allele; one Siberian husky and three Alaskan sled dogs were identified as carriers. The affected chromosomes from the AM, MAS, and Siberian husky were genotyped for 147 SNPs in a 3.93 Mb interval within the cd locus. An identical shared affected haplotype, 0.5 Mb long, was observed in all three breeds and defined the minimal linkage disequilibrium (LD) across breeds. This supports the idea that the mutated allele was identical by descent (IBD). Conclusion We report the occurrence of CNGB3-achromatopsia in a new canine breed, the MAS. The CNGB3-deletion allele previously described in the AM was also observed in a homozygous state in the affected MAS, as well as in a heterozygous carrier state in a Siberian husky and Alaskan sled dogs. All affected alleles were shown to be IBD, strongly suggesting an affected founder effect. Since the MAS is not known to be genetically related to the AM, other breeds may potentially carry the same cd-allele and be affected by achromatopsia.

Background Two strains of the silver fox (Vulpes vulpes), with markedly different behavioral phenotypes, have been developed by long-term selection for behavior. Foxes from the tame strain exhibit friendly behavior towards humans, paralleling the sociability of canine puppies, whereas foxes from the aggressive strain are defensive and exhibit aggression to humans. To understand the genetic differences underlying these behavioral phenotypes fox-specific genomic resources are needed. Results cDNA from mRNA from pre-frontal cortex of a tame and an aggressive fox was sequenced using the Roche 454 FLX Titanium platform (> 2.5 million reads & 0.9 Gbase of tame fox sequence; >3.3 million reads & 1.2 Gbase of aggressive fox sequence). Over 80% of the fox reads were assembled into contigs. Mapping fox reads against the fox transcriptome assembly and the dog genome identified over 30,000 high confidence fox-specific SNPs. Fox transcripts for approximately 14,000 genes were identified using SwissProt and the dog RefSeq databases. An at least 2-fold expression difference between the two samples (p < 0.05) was observed for 335 genes, fewer than 3% of the total number of genes identified in the fox transcriptome. Conclusions Transcriptome sequencing significantly expanded genomic resources available for the fox, a species without a sequenced genome. In a very cost efficient manner this yielded a large number of fox-specific SNP markers for genetic studies and provided significant insights into the gene expression profile of the fox pre-frontal cortex; expression differences between the two fox samples; and a catalogue of potentially important gene-specific sequence variants. This result demonstrates the utility of this approach for developing genomic resources in species with limited genomic information.

The silver fox provides a rich resource for investigating the genetics of behavior, with strains developed by intensely selective breeding that display markedly different behavioral phenotypes. Until recently, however, the tools for conducting molecular genetic investigations in this species were very limited. In this review, the history of development of this resource and the tools to exploit it are described. Although the focus is on the genetics of domestication in the silver fox, there is a broader context. In particular, one expectation of the silver fox research is that it will be synergistic with studies in other species, including humans, to yield a more comprehensive understanding of the molecular mechanisms and evolution of a wider range of social cognitive behaviors.

The foxes at Novosibirsk, Russia, are the only population of domesticated foxes in the world. These domesticated foxes originated from farm-bred silver foxes (Vulpes vulpes), whose genetic source is unknown. In this study we examined the origin of the domesticated strain of foxes and two other farm-bred fox populations (aggressive and unselected) maintained in Novosibirsk. To identify the phylogenetic origin of these populations we sequenced two regions of mtDNA, cytochrome b and D-loop, from 24 Novosibirsk foxes (8 foxes from each population) and compared them with corresponding sequences of native red foxes from Europe, Asia, Alaska and Western Canada, Eastern Canada, and the Western Mountains of the USA. We identified seven cytochrome b - D-loop haplotypes in Novosibirsk populations, four of which were previously observed in Eastern North America. The three remaining haplotypes differed by one or two base change from the most common haplotype in Eastern Canada. Φ(ST) analysis showed significant differentiation between Novosibirsk populations and red fox populations from all geographic regions except Eastern Canada. No haplotypes of Eurasian origin were identified in the Novosibirsk populations. These results are consistent with historical records indicating that the original breeding stock of farm-bred foxes originated from Prince Edward Island, Canada. Mitochondrial DNA data together with historical records indicate two stages in the selection of domesticated foxes: the first includes captive breeding for ~50 years with unconscious selection for behaviour; the second corresponds to over 50 further years of intensive selection for tame behaviour.

Directional asymmetry (DA) is a characteristic of most vertebrates, most strikingly exhibited by the placement of various organs (heart, lungs, liver, etc.) but also noted in small differences in the metrics of skeletal structures such as the pelvis of certain fish or sauropsids. We have analyzed DA in the skeleton of the fox (V. vulpes), using ~1,000 radiographs of foxes from populations used in the genetic analysis of behavior and morphology. Careful measurements from this robust data base demonstrate that: 1) DA occurs in the limb bones, the ileum, and ischium and in the mandible; 2) regardless of the direction of the length asymmetry vector of a particular skeletal unit, the vectorial direction of length is always opposite to that of width; 3) with the exception of the humerus and radius, there is no correlation or inverse correlation between vectorial amplitudes or magnitudes of bone asymmetries. 4) Postnatal measurements on foxes demonstrate that the asymmetry increases after birth and continues to change (increasing or decreasing) during postnatal growth. 5) A behavior test for preferential use of a specific forelimb exhibited fluctuating asymmetry but not DA. None of the skeletal asymmetries were significantly correlated with a preferential use of a specific forelimb. We suggest that for the majority of fox skeletal parameters, growth on the right and left side of the fox are differentially biased resulting in fixed differences between the two sides in either the rate of growth or the length of the period during which growth occurs. Random effects around these fixed differences perturb the magnitude of the effects such that the magnitudes of length and width asymmetries are not inversely correlated at the level of individual animals.

During the second part of the twentieth century, Belyaev selected tame and aggressive foxes (Vulpes vulpes), in an effort known as the “farm-fox experiment”, to recapitulate the process of animal domestication. Using these tame and aggressive foxes as founders of segregant backcross and intercross populations we have employed interval mapping to identify a locus for tame behavior on fox chromosome VVU12. This locus is orthologous to, and therefore validates, a genomic region recently implicated in canine domestication. The tame versus aggressive behavioral phenotype was characterized as the first principal component (PC) of a PC matrix made up of many distinct behavioral traits (e.g. wags tail; comes to the front of the cage; allows head to be touched; holds observer’s hand with its mouth; etc.). Mean values of this PC for F1, backcross and intercross populations defined a linear gradient of heritable behavior ranging from tame to aggressive. The second PC did not follow such a gradient, but also mapped to VVU12, and distinguished between active and passive behaviors. These data suggest that (1) there are at least two VVU12 loci associated with behavior; (2) expression of these loci is dependent on interactions with other parts of the genome (the genome context) and therefore varies from one crossbred population to another depending on the individual parents that participated in the cross.

Rod-cone dysplasia type 2 (rcd2) is an autosomal recessive disorder that segregates in collie dogs. Linkage disequilibrium and meiotic linkage mapping were combined to take advantage of population structure within this breed and to fine map rcd2 to a 230-kb candidate region that included the gene C1orf36 responsible for human and murine rd3, and within which all affected dogs were homozygous for one haplotype. In one of three identified canine retinal RD3 splice variants, an insertion was found that cosegregates with rcd2 and is predicted to alter the last 61 codons of the normal open reading frame and further extend the open reading frame. Thus, combined meiotic linkage and LD mapping within a single canine breed can yield critical reduction of the disease interval when appropriate advantage is taken of within-breed population structure. This should permit a similar approach to tackle other hereditary traits that segregate in single closed populations.

The features of modern dog breeds that increase the ease of mapping common diseases, such as reduced heterogeneity and extensive linkage disequilibrium, may also increase the difficulty associated with fine mapping and identifying causative mutations. One way to address this problem is by combining data from multiple breeds segregating the same trait after initial linkage has been determined. The multibreed approach increases the number of potentially informative recombination events and reduces the size of the critical haplotype by taking advantage of shortened linkage disequilibrium distances found across breeds. In order to identify breeds that likely share a trait inherited from the same ancestral source, we have used cluster analysis to divide 132 breeds of dog into five primary breed groups. We then use the multibreed approach to fine-map Collie eye anomaly (cea), a complex disorder of ocular development that was initially mapped to a 3.9-cM region on canine chromosome 37. Combined genotypes from affected individuals from four breeds of a single breed group significantly narrowed the candidate gene region to a 103-kb interval spanning only four genes. Sequence analysis revealed that all affected dogs share a homozygous deletion of 7.8 kb in the NHEJ1 gene. This intronic deletion spans a highly conserved binding domain to which several developmentally important proteins bind. This work both establishes that the primary cea mutation arose as a single disease allele in a common ancestor of herding breeds as well as highlights the value of comparative population analysis for refining regions of linkage.