ILAR Journal Online, Volume 39(2/3) 1998: Comparative Gene Mapping

Online Issues

<< All Back-issues

<< This Issue's Table of Contents

ILAR Journal V39(2/3) 1998
Comparative Gene Mapping

The Fox Gene Map
Nikolai B. Rubtsov

Nikolai B. Rubtsov, Ph.D., is with the Institute of Cytology and Genetics, Academy of Sciences of Russia, Siberian Department, Novosibirsk, Russia.

INTRODUCTION

The wild type red fox is common to Europe, Asia, North Africa, and North America. Furthermore, it has accommodated itself so well in Australia, where it was taken from England, that it has spread over the entire continent. It adapts easily to different climates and biocoenoses--in the open; in the forest, highland or lowland; on the coast; or inland. In 1827, C. A. Bjorkman was the first to try to make the cross fox reproduce in captivity, but his attempt failed. Modern fur animal farming is believed to have been started in 1895 on Prince Edward Island, St. Lawrence Bay, Canada, by Charles Dalton, Robert Oulton, and Robert Rayner. Their animals were the origin of the farm-bred standard silver fox. Another stock of silver fox was associated with animals bred by J. Beetz, a pair of which was shipped to Norway in 1914. Over the past 100 yr, fur animal breeding has developed into an industry of great economic importance, especially in Nordic countries.

Inheritance of Color Types

The silver fox was first regarded as a mutation of the red fox. It was thought to belong to a minor group; however, the export of hundreds of silver fox skins in former centuries from North America implies that the silver color type was common to many populations. At least 2 black mutants were known: the Canadian, from which the standard silver fox originates; and the Alaskan silver fox, which is larger, with brown hairs in and around the ears and a coarser pelt with a brownish tint. The amount of silver-bar hairs is under polygenic control in both. Guard hairs with silver bars are also present in the red fox, whose coat color differs from population to population. Warwick and Hanson (Ashbrook 1937) hypothesized that genes A and B and their allelic forms a and b are responsible for the silver and Alaskan fox color types. Incomplete dominance of A and B over a and b, respectively, has been demonstrated. The red fox (AABB) is considered to be Vulpes vulpes wild type, the AAbb combination is the standard silver fox, and aaBB is the Alaskan silver fox. The other genotypes correspond to the substandard silver fox, sub-Alaskan silver fox, double silver fox, gold fox (smoky red), cross fox, and blended cross fox. The features of spine stripe formation led Ilina (Ness and others 1988) to propose that the spine stripe must be dependent on a separate factor (D). Since the amount of red color and spine stripe clearness are subject to strong variation, we do not yet understand the genetic control of the black or red color in the fox. Many other colors have been established through years of breeding and selection. Originally the mutant alleles were largely confined to the silver fox. Later, they were introduced in various combinations to red and cross fox types. Only recently have the mutations been listed and described (Ness and others 1988).

Fox Karyotype

The fox chromosomes were first described by Wodsedalek in 1931, when he reported 42 as the chromosome number of the male red fox. Later, in 1938, Andres found the number to be 34. In 1942, Wipf and Scackelford described a fox karyotype of 34 chromosomes of various lengths, including a pair of satellite chromosomes (Wipf and Scackelford 1942). Actually, the chromosome complement of the red fox has 32 metacentric autosomes; the metacentric X chromosome; the acrocentric Y chromosome; and a few, from 0 to 8, B chromosomes. The banding patterns of fox chromosomes make it possible to easily identify all the fox chromosomes (Graphodatsky and Radjabli 1981; Mäkinen 1985). More recently, higher resolution banding has led to the nomenclature of 510 bands per haploid set (Graphodatsky and others 1995) (Figure 1). Nucleolus organizer regions (NORs¹)are located in the telomeric regions of q-arms of chromosomes 8, 9, and 13 (Mäkinen 1985; Graphodatsky and Radjabli, 1988) (Figure 1).

Fox B chromosomes are very small, as is the Y chromosome. They were first described by Gustavsson and Sundt (1967), 2 yr after the first reported discovery of mammalian B chromosomes found in the greater glider Shoinobates volans (now designated Petauroides volans) (Hayman and Martin 1965). Morphologically, fox B chromosomes can be uni- or biarmed, although their small size often prevents an accurate description (Belyaev and others 1974a). Gustavsson and Sundt (1967) suggested that B chromosomes are heterochromatic, genetically inert chromosomes. Volobujev and others (1976) reported that B chromosomes replicate during the second half of S phase and terminate replication at the same time as late-replicated regions of other chromosomes. The somatic B chromosomes are often smaller than 1 mm, and high-resolution techniques do not lead to their elongation in prometaphase spreads. Staining with Giemsa with trypsin (GTG¹)and R-bands induced by pretreatment synchronized cultures with BrdU and subsequent staining with acridine orange (RBA¹)techniques has revealed no banding patterns (Switonsky and others 1987). Nevertheless, RBA-stained B chromosomes can be distinguished from the Y chromosome because they stain less brightly, indicating that the B chromosomes have late-replicating DNA. It should be mentioned, however, that Ellenton and Basrur (1980) reported replication of fox B chromosomes at mid-S phase. They argued that the process is probably complete before replication commences in the late-replicated chromosomal regions. Results of their investigations of mitotic chromosomes and the staining properties of B chromosomes at pachytene indicated that the B chromosomes were likely to contain both heterochromatin and euchromatin (Ellenton and Basrur, 1980). Low and Benirschke (1972) hypothesized also that B chromosomes could be distal fragments of the chromosomes of the basic set, although others who have observed the B chromosomes reject this hypothesis (Renzoni and Omodeo 1972). Although the B chromosomes sometimes vary in morphology as observed by light microscopy (Belyaev and others 1974a), analysis of the synaptonemal complex of the B chromosomes has shown identical kinetochore location (Switonsky and others 1987). Pairing behavior of B chromosomes in meiosis is also indicative of their homology. Two B chromosomes occurring in the same cell are normally paired as a bivalent with 1 central and 2 lateral elements, although 2 univalents are sometimes observed. As univalents, the B chromosomes demonstrate a progressively folding-back behavior, ending up in intrachromosome pairing. Three B chromosomes in the same cell mainly become trivalent but sometimes form 2 normally paired chromosomes and 1 folded univalent. Foldback configuration in meiosis indicates the presence in the B chromosomes of repeated DNA sequences. Some repeats, common to heterochromatic regions of all silver fox chromosomes and heterochromatic arms of blue fox chromosomes, were also found in the heterochromatic regions of B chromosomes (A. S. Graphodatsky, personal communication). As mentioned above, the number of B chromosomes varies from animal to animal. Moreover, variation in B chromosome number among the cells of the same animal has been found, a phenomenon that has been studied by chromosome analysis of hundreds of foxes. Two types of foxes have been described. In some animals, the number of B chromosomes is identical in all cells. In other animals (mosaics), the number varies among the cells of the same individual due to the existence of different cell clones (Belyaev and others 1974a). The number of B chromosomes is independent of somatic tissues or seasons, but the mean number of B chromosomes in reproductive tissue cells was found to be higher than that in somatic cells (Radjabli and others 1978).

Despite all studies, the data on the genetic role of the fox B chromosomes and mammalian B chromosomes in general are scanty. A series of investigations (Belyaev and others 1974a,b; Volobujev and Radjabli 1974; Volobujev and others 1976) was carried out on the silver foxes to detect the effect of the B chromosomes. The B chromosome variation patterns were analyzed in the groups of foxes with different, genetically determined types of behavior (Belyaev and others 1974b). The first group consisted of the foxes selected for placid behavior toward humans over 10 generations; the second group was composed of the foxes selected for aggressiveness over 3 to 5 generations; and the third group consisted of the animals bred under usual farming conditions. The frequency of mosaics was twice as high in the groups of foxes selected for behavior as in the third group. Shell-hammer (1969) suggested also that B chromosomes might influence physiological characteristics--in particular, behavior. It is difficult to discuss the nature of this influence, but it should be noted that the results mentioned above do not support the idea of the genetic inertness of the B chromosomes, at least in foxes. Nevertheless, the fox B chromosomes are still largely considered remnants of ancient chromosomes involved in numerous structural rearrangements during karyotypic evolution. The distribution of chromosome numbers in the Canidae provides a good basis for this hypothesis. Probably we will find answers to the many remaining questions only through generation of the B chromosome-specific DNA libraries and analysis of DNA sequences specific for B chromosomes.

REASONS FOR MAPPING THIS SPECIES

Comparative Gene Mapping

The silver fox belongs to a large family of dog-like carnivores, the Canidae, which includes 36 living species of 16 genera (Nowak and Paradiso 1983). Canidae stand out from the other carnivore families. Carnivora comprise 7 families or 2 superfamilies, Canoidea and Feloidea. Chromosome morphology and GTG banding patterns in Feloidea species are mostly conserved. For example, most chromosome pairs (15 of 18 or 19) were found cytologically invariant among 37 Felidae species (Wayne and others 1987a,b; Wurster-Hill and Centerwall 1982). Chromosomes with identical or similar GTG banding patterns have also been discovered in Hyaenidae, Viverridae, Procyonidae, and Mustelidae. In Canidea, the karyotypes are extremely variable with respect to both chromosome number and morphology (Graphodatsky 1989; Wayne and others 1987a). The chromosome number ranges from 2n = 34 plus a few B chromosomes in the red fox (Vulpes vulpes) to 2n = 78 in the domestic dog (Canis familiaris) and the gray wolf (Cants lupus). Nevertheless, comparison of the GTG banding patterns of chromosomes in canids revealed extended regions of homology covering almost all their chromosomes (Graphodatsky 1989; Wurster-Hill and Centerwall 1982). At the same time, the chromosomes and chromosome regions in Canidea are strongly different from those in Feloidea. Extensive cytogenetic and linkage/ synteny homologies observed on the cat, mink, and even human chromosomes (Nash and O'Brien, 1982; Rubtsov and others 1988) have underscored the importance of fox gene mapping and of thoroughly comparing fox chromosomes with those of the cat, mink, and human.

Representation of Fur-bearing Animals

Another incentive for fox gene mapping is recognition of the silver fox as one of the most important species in the fur industry. Fox breeders have long been searching for and identifying mutations that affect coat color. Recently, the results have been described in detail and summarized in Beautiful Fur Animals and Their Color Genetics (Ness and others 1988). Despite many coat color mutations identified in the fox, no direct evidence exists for chromosomal localization of the genes responsible for these mutations.

Experimental Domestication of Animals

Thirty years ago, the silver fox was chosen as a model for experimental domestication of animals. On the one hand, the lox is closely related to the dog, which represents the pinnacle of evolutionary transformations achieved by domestication. On the other hand, the fox has great ecological flexibility, which allows it to adapt to a wide range of conditions. Large-scale experiments were started at the beginning of the 1960s with the aim of establishing a population of foxes that would resemble dogs in behavior. The population has now passed through more than 30 successive generations of strict selection, and the foxes behave very much as dogs behave. The dog-like foxes follow their master, licking and begging for favors. The tame foxes also have complex physiological and morphological distinctions. The reproductive function tends to a weaker dependence on season, and the mean ovulation rate has changed. Morphological novelties not characteristic of the fox as a species have been observed. The results of these investigations are summarized in Evolutionary-Genetic and Genetic-Physiological Aspects of Fur Animal Domestication--A Collection of Reports (Trut and Osadchuk 1997).

APPROACHES USED TO DEVELOP THE MAP

Somatic Cell Hybridization

Somatic cell hybridization has proven to be an efficient technique for fox gene mapping. This approach requires somatic hybrids with appropriate chromosome segregation, reliable identification of chromosomes in hybrid cells, and a low level of chromosome rearrangements (Rubtsov and others 1981, 1987). The problems that may arise from unidentified chromosome rearrangements have been discussed in detail elsewhere (Cowmeadow and Ruddle 1978; Rubtsov and others 1981; Wijnen and others 1977). To avoid errors in fox gene assignments, special criteria were taken into account (Rubtsov and others 1981, 1987, 1988). More than 200 fox-hamster independent somatic hybrids were obtained. The chromosome contents of the hybrids were estimated by the electrophoretic patterns of some fox enzymes, and 42 hybrids were selected as having the combinations of fox chromosomes most useful for study. The clones were karyotyped, and those carrying rearranged fox chromosomes were discarded. The cells of the remaining 23 clones were propagated up to a number of 10⁸and karyotyped again.

To increase the reliability of the clone panel, a chromosome analysis of the clones was performed again, this time using a high-resolution GTG-banding technique (Rubtsov and others 1987; 1988). Notably, assignment requires not only a high level of concordant segregation of the gene and the chromosome but also a sufficiently high level of discordant segregation of the gene and every other chromosome. All hypotheses of an association between the gene and any other chromosome were disproved (Cowmeadow and Ruddle 1978; Rubtsov and others 1981; Wijnen and others 1977). The panel was designed as a set of hybrids with at least 6 clones in which any pair of the fox chromosomes displayed discordant segregation. The minimum percentage of clones displaying discordance for any chromosome pair was more than 25% (not less than 6 of 23 hybrids) (Rubtsov and others 1987, 1988). The panel thus became a tool for assigning to a particular chromosome any fox gene with detectable presence in the fox-hamster somatic hybrids.

The chromosomal distribution of 35 fox genes also has been determined using the panel clones (Adkison and others 1994; Koroleva and others 1996; Nesterova and others 1991a,b, 1992; Rubtsov and others 1988). These studies either analyzed the expression of fox isozymes or applied Southern blot hybridization with appropriate DNA probes. At the time of this writing, 15 autosomes and the X chromosome have been marked. To date, no gene has been assigned to fox chromosome 13 or to the Y chromosome. The results of fox gene mapping are summarized in Figure 1.

Gene Linkage Data

Despite the large number of coat color mutations identified in the fox, the only information on genetic linkage available to date is that the factor D and the Alaskan gene are probably linked (Ness and others 1988). The development of the fox linkage map is an important goal for further exploiting the fox in comparative gene mapping studies and for further developing its economic potential. Heterologous DNA probes can be used for this purpose, as has been done to identify gene linkages in the dog (Sack and others 1996; also see Binns and others 1998).

Intergeneric Fox Hybrids (Alopex lagopus x Vulpes vulpes)

An enzyme comprising 2 or more subunits encoded by the same gene may exhibit several isozymes in animals that are heterozygous for the gene. In addition to the isozymes observed in the respective homozygotes, 1 or more isozymes in cells of heterozygotes consist of a combination of subunits encoded by different alleles. However, there is no hybrid isozyme(s) if the gene is located on the X chromosome, due to inactivation of 1 of the X chromosomes in somatic cells. Therefore, for dimeric or multimeric isozymes, a pattern without hybrid isozyme(s) should be regarded as an indica-tion of X-chromosomal localization of the gene. G6PD is a dimeric enzyme in mammals. It was shown that although blue and silver lox G6PD have different electrophoretic mobilities, no hybrid isozyme was observed in Alopex lagopus x Vulpes vulpes (Serov and others 1978). These data indicate the X-chromosomal localization of the G6PD gene. G6PD is the only example of gene assignment by intergeneric fox hybrids.

Cytological Visualization of Gene Location

The cytological technique developed for detection of active NORs makes it possible to map genes for rRNA (Howell 1977). Active NORs were thus mapped on fox chromosomes 8, 9, and 13 (Graphodatsky and Radjabli 1988; Mäkinen 1985) (Figure 1).

Refinement of in situ hybridization techniques for gene mapping has led to the localization of a large number of genes in humans and other species. The banding patterns of the fox chromosomes make them easily identifiable, so fox gene mapping by in situ hybridization depends on the avail-ability of appropriate DNA probes. Some DNA probes for human genes have been used in an attempt to localize the homologous lox genes. Unfortunately, the only success was achieved with the probe for human esterase D, which made possible the assignment of its fox homologue to 6q(cen-->q 17) (Graphodatsky and others 1991 ). Neverthe-less, the results of dog gene mapping by fluorescence in situ hybridization (FISH¹)with human yeast artificial chromo-some clones as well as genomic clones of canine origin (Dutra and others 1996) showed that the modern technique of FISH can probably be used for mapping fox loci.

CURRENT MAP STATUS

Unidentified chromosome rearrangements and different sen-sitivities of the methods used for detection of the gene marker and chromosome may be responsible for erroneous gene as-signments by the somatic cell hybridization technique. The following categories of certainty have been recommended (Ruddle and Giblett 1975): confirmed (C) or established (at least 2 groups of investigators agree); provisional (P) or un-confirmed (data from 1 group only); inconsistent (1) or con-tradictory (groups disagree).

Segregation analysis of gene markers and fox chromo-somes suggests that the inferences of all gene assignments are very reliable. Nevertheless, because all gene assignments except that of the ESD gene were performed using the only existing panel of fox-hamster somatic hybrids, the assignments should be categorized as provisional.

With regard to the ESD gene, independent data were obtained by in situ hybridization of the DNA probe for human ESD on fox chromosomes (Graphodatsky and others 1991). The gene was located on fox chromosome 6 (cen--->q 17), and therefore the gene assignment is confirmed.

To determine the reliability of the fox gene assignments, all results obtained from the fox-hamster hybrid clone panel were reviewed retrospectively. A total of 35 fox genes had been assigned using the panel. Among 805 cases of gene-chromosome segregation, only 8 cases of "false" discordance were detected (less than 1% of discordance), implying that the lox-hamster panel is a powerful and reliable tool for mapping fox genes. Furthermore, to minimize segregation data errors, the sensitivities of the methods for detection of gene markers were estimated. The amount of gene product and the number of cells with the fox chromosome were taken into account. For all the genes, a good correlation was observed between the number of cells with a particular fox chromosome and the amount of gene product (Rubtsov and others 1987). Although these data do not change the status of the fox gene assignments from provisional, they do indicate that the chromosomal assignments are highly reliable.

SCIENTIFIC CONTRIBUTIONS OF THE MAP

Comparative chromosome-banding studies on many mammalian species have revealed extended chromosome regions of banding pattern homology within an order. Furthermore, homologous regions have been found on the chromosomes of the cat, mink, human, and some other mammals.

The canids were not studied for many years. No gene assignments had been made, and the banding patterns of canid chromosomes were known to differ drastically even from the chromosomes of Feloidea species. Now it is possible to use the fox in comparative studies of mammalian chromosomes because some gene assignments have been made and because some regions of banding homology have been detected between fox chromosomes and mink and cat chromosomes. Some examples are shown in Figure 2.

Studies of karyotype evolution in carnivores based on the data from fox, mink, and cat gene mapping provide evidence for a stage of intensive karyotype evolution followed by the formation of the canid branch of the carnivore phylogenetic tree. The chromosome rearrangements that led to canid chromosomes were associated with multiple disruptions of ancient groups of syntenic genes. Therefore, chromosome rearrangements in the canids were not restricted to the shuffling of material within the same chromosome, as has been demonstrated on some bovine chromosomes (Hayes 1995). Combined data suggest that at the earlier stages of formation of some mammalian taxa, extensive transformations of the original karyotype took place after which new chromosome elements and groups of linkage genes formed. Upon completion of that stage, karyotype evolution within the phylum probably continued due to recombination of the new basic elements and variations in heterochromatic regions (Rubtsov and others 1988; Graphodatsky 1989).

USES OF THE MAP AND ACCESSIBILITY

The fox gene map provides an excellent resource for further study of karyotype evolution in Carnivora. Initially this resource can help identify subchromosomal homologous regions. Subsequent gene mapping can independently verify hypotheses from comparative cytogenetic studies. In the future, comparative gene mapping in combination with the visualization of the homologous regions of chromosomes by region-specific DNA libraries is expected to be especially useful. Studies of other species, employing chromosome-specific painting probes, are in progress (Ferguson-Smith and others 1998; Glas and others 1998; Toder and others 1998; Wienberg and Stanyon 1998).

Despite rapid progress in canine genetics, some problems remain with dog chromosome nomenclature and reliable identification of all dog chromosomes. Comparative GTG-banding analysis indicates a close relationship between some chromosome arms in the silver fox and dog chromosomes (Graphodatsky 1989; Wurster-Hill and Centerwall 1982). The inferred homologous relationships can be checked by generation of fox chromosome arm-specific DNA libraries by chromosome microdissection and ZOO-FISH of these libraries as probes on dog chromosomes. The efficiency of this approach has been demonstrated in studies with microdissected arm-specific paints for other species (Ponce de León and others 1996; Rubtsov and others 1997; Chaudhary and others 1998). Reliable detection of homology between particular fox chromosome arms and dog chromosomes will enable fox chromosomes to be used as a convenient control system for checking gene assignments in the dog.

CONCLUSION

Comparisons of chromosome polymorphisms in Carnivora reveal a surprising conservatism of chromosome arms within a family. The characteristic karyotypes within each family were formed as different combinations of the basic elements of the set. Species of the Felidae family have stable, virtually identical chromosome complements. The chromosomes of mustelids, except skunk, consist mainly of large homologous regions and blocks of heterochromatin. Despite considerable variation in the number and morphology of the canid chromosomes, the same applies to Canidae. Studies of karyotype evolution in mammals should include a comparison of the basic elements. Detection of homologous regions requires wide use of markers of homology in addition to cytological techniques.

Although the fox gene map is rudimentary, it is now at a stage where it is useful in comparative gene map analyses of mammalian chromosomal evolution.

¹Abbreviations used in this paper: FISH, fluorescence in situ hybridization; GTG, Giemsa banding with trypsin; NOR, nucleolus organizer region; RBA, R-bands induced by pretreatment synchronized cultures with BrdU and subsequent staining with acridine orange.

ACKNOWLEDGMENTS

I thank V. Filonenko for help with the English language and Dr. Alexander S. Graphodatsky for permission to cite unpublished data.

REFERENCES

Adkison LR, Koroleva IV, Zakian SM. 1994. Mapping of the *-subunit of the fox fibronectin receptor (FNRA) to chromosome 8. Mamm Genome 5:119-120.

Ashbrook FD. 1937. The breeding for fur animals. Yearbook of Agriculture (US Department of Agriculture) Washington DC: US GPO. p 1379-1395.

Belyaev DK, Volobuev VT, Radjabli SI, Trut LN. 1974a. Polymorphism and mosaicism for additional chromosomes in silver foxes. Genetika 1(1:58-67.

Belyaev DK, Volobuev VT, Radjabli SI, Trut LN. 1974b. Investigation of the nature and the role of additional chromosomes in silver fox. II. Additional chromosomes and breeding of animals for behaviour. Genetika 10:83-91.

Binns M, Holmes N, Breen M. 1998. The dog gene map. ILAR J 39:177-181.

Chaudhary R, Raudsepp T, Guan XY, Zhang H, Chowdhary BP. 1998. Zoo-FISH with microdissected arm specific paints for HSA2, 5, 6, 16, and 19 refines known homology with pig and horse chromosomes. Mamm Genome 9:44-49.

Cowmeadow MP, Ruddle FH. 1978. Computer-assisted statistical procedures for somatic gene assignment. Cytogenet Cell Genet 22:694-697.

Dutra AS, Mignot E Puck JM. 1996. Gene localization and syntenic mapping by FISH in the dog. Cytogenet Cell Genet 74:113-117.

Ellenton JA, Basrur PK. 1980. Microchromosomes of the Ontario red fox (Vulpes vulpes): An attempt at characterization. Can J Genet Cytol 22:553-567.

Ferguson-Smith MA, Yang F, O'Brien PCM. 1998. Comparative mapping using chromosome sorting and painting. ILAR J 39:68-76.

Glas R, Wakefield MJ, Toder R, Graves JAM. 1998. Comparative chromosome painting maps. ILAR J 39 (Poster).

Graphodatsky AS. 1989. Conserved and variable elements of mammalian chromosomes. In: Halnan CRE, editor. Cytogenetics of Animals. Tucson AZ: CAB International North America. p 95-123.

Graphodatsky AS, Beklemisheva VR, Doll G. 1995. High-resolution GTG-banding patterns of dog and silver fox chromosomes: Description and comparative analysis. Cytogenet Cell Genet 69:226-231.

Graphodatsky A, Lushnikova T, Biltueva L, Eremina V, Rubtsov N. 1991. Localization of some human genes to mammalian chromosomes by in situ hybridisation. Cytogenet Cell Genet 58:51.

Graphodatsky AS, Radjabli SI. 1981. Comparative cytogenetics of three canids species (Carnivora, Canidae). II. Distribution of C-heterochromatin. Genetika 17:1500-1504.

Graphodatsky AS, Radjabli SI. 1988. The Chromosomes of Agricultural and Laboratory Mammals. All Atlas. Novosibirsk: Nauka. p 1-128.

Gustavsson 1, Sundt CO. 1967. Chromosome elimination in the evolution of the silver fox. J Hered 58:75-78.

Hayes H. 1995. Chromosome painting with human chromosome-specific DNA libraries reveals the extent and distribution of conserved segments in bovine chromosomes. Cytogenet Cell Genet 71:168-174.

Hayman DL, Martin PG. 1965. Supernumerary chromosomes in the marsupial Shoinobates volans (Kerr). Aust J Biol Sci 18:1081 - 1082.

Howell WM. 1977. Visualization of ribosomal gene activity: Silver stains proteins associated with rRNA transcribed from oocyte chromosomes. Chromosoma 62:361-367.

Koroleva IV, Malchenko SN, Brusgaard K, Khlebodarova TM, Rubtsov N, Zakian SM. 1996. Chromosome localization of the genes for growth hormone, somatostatin peptide, ornithine transcarbamylase, and prion protein in silver fox (Vulpes fulvus). Mamm Genome 7:860-862.

Low R J, Benirschke K. 1972. Microchromosomes in the American red fox (Vulpes fulva). Cytologia 37:1-11.

Mäkinen A. 1985. The standard karyotype of the silver fox (Vulpes fulvus Desm). Committee for the standard karyotype of Vulpes fulvus Desm. Hereditas 103:289-297.

Nash WG, O'Brien SJ. 1982. Conserved regions of homologous G-banding chromosomes between orders in mammalian evolution: Carnivore and primates. Proc Natl Acad Sci U S A 79:6631-6635.

Ness NN, Einarson EJ, Lohi O, Jorgensen G. 1988. Beautiful Fur Animals and Their Color Genetics. Glostrup Denmark: Scientifur.

Nesterova T, Mazurok N, Nikitina I, Matveeva V, Rubtsov N, Zakian S. 1992. Gene map of the silver fox (Vulpes fulvus). In: O'Brien S J, editor. Genetic Maps. Cold Spring Harbor NY: Cold Spring Harbor Press. p 4.256-4.257.

Nesterova T, Nikitina 1, Zakian S, Rubtsov N, Matveeva V, Radjabli S. 1991a. Mapping of the silver fox genes: Chromosomal localization of the genes for GOT2, AK1, ALDOC, ACP1, ITPA, PGP, and BLVR. Cytogenet Cell Genet 56:185-188.

Nesterova T, Rubtsov N, Zakian S, Matveeva V, Graphodatsky A. 1991b. Mapping of the silver fox genes: Assignments of the genes for ME1, ADK, PP, PEPA, GSR, MPI, and GOT1. Cytogenet Cell Genet 56:125-127.

Nowak RM. Paradiso JL. 1983. Walker's Mammals of the World, 4th edition. Baltimore MD: Johns Hopkins University Press.

Ponce de León FA, Ambady S, Hawkins GA. Kappes SM, Bishop MD, Robl JM, Beattie CW. 1996. Development of a bovine X chromosome linkage group and painting probes to assess cattle, sheep, and goat X chromosome segment homologies. Proc Natl Acad Sci U S A 93:345(I-3454.

Radjabli SI, lsaenko AA, Volobujev VT. 1978. Investigation of the nature and the role of the additional chromosomes in silver foxes. IV. B chromosome behavior in meiosis. Genetika 14:438-443.

Renzoni A, Omodeo P. 1972. Polymorphic chromosome system in the fox. Caryologia 25:173-187.

Rubtsov N, Graphodatsky A, Matveeva V, Radjabli S, Nesterova T, Kulbakina N, Zakian S. 1988. Silver fox gene mapping: Conserved chromosome regions in the order Carnivora. Cytogenet Cell Genet 48:95-98.

Rubtsov N, Matveeva V, Radjabli S, Kulbakina N, Nesterova T, Zakian S. 1987. Construction of a clone panel of fox-Chinese hamster somatic cell hybrids and assignment of genes for LDHA, LDHB, GPI, ESD, G6PD, HPRT, GALA in the silver fox. Genetika 23:1088-1096.

Rubtsov NB, Radjabli SI, Gradov AA, Serov OL. 1981. Chinese hamster x American mink somatic cell hybrids: Characterization of a clone panel and assignment of the mink genes for malate dehydrogenase, NADP-1 and malate dehydrogenase, NAD-1. Theor Appl Genet 60:99-106.

Rubtsov N, Serdyukova N, Kaftanovskaya E, Yang F, Biltueva L, Vorobieva N, Rogacheva M, Oda S, Graphodatsky A. 1997. Visualization of the cattle Xp homologous regions on the X chromosomes of some pecorans by chromosome microdissection and heterologous painting. Cytology 62:203-208.

Ruddle FH, Giblett ER. 1975. Report of the committee (m the genetic constitution of autosomes other than I and 2. Cytogenet Cell Genet 14:183-189.

Sack GH Jr, Taylor EW, Meyers DA, Dragwa CR, Cork LC. 1996. Canine genetic linkage study using heterologous DNA probes. J Hered 87:15-20.

Serov OL, Zakijan SM, Kulichkov VA. 1978. Allelic expression in intergeneric lox hybrids (Alopex lagopus x Vulpes vulpes). 111. Regulation of the expression of the parental alleles at the Gpd locus linked to thc X-chromosome. Biochem Genet 16:145-157.

Shellhammer HS. 1969. Supernumerary chromosomes of the harvest mouse, Reithrodontomys megalotis. Chromosoma 27:1 I12-108.

Switonsky M, Gustavsson I, Hoejer K, Ploeen L. 1987. Synaptonemal complex analysis of the B chromosomes in spermatocytes of the silver fox (Vulpes fulvus Desm.). Cytogenet Cell Genet 45:84 92.

Toder R, O'Neill RJW, Graves JAM. 1998. Chromosome painting in marsupials. ILAR J 39:92-95.

Trut LN, Osadchuk LV, editors. Evolutionary-Genetic and Genetic-Physiological Aspects of Fur Animal Domestication--A Collection of Reports. IFASA/SCIENTIFUR 1997. Oslo: Oslu Fur Centre.

Volobujev VT, Radjabli SI. 1974. Investigation of the nature and the role of the additional chromosomes in silver foxes. I. Comparative analysis of additional chromosomes in different tissues and type preparations and at different seasons. Genetika 10:77-82.

Volobujev VT, Radjabli SI, Belyaeva ES. 1976. Investigation of the nature and the role of the additional chromosomes in silver foxes. Ill. Replication pattern in additional chromosomes. Genetika 12:311-34.

Wayne RK, Nash WG, O'Brien SJ. 1987a. Chromosome evolution of the Canidae. I. Species with high diploid number. Cytogenet Cell Genet 44: 123-133.

Wayne RK, Nash WG, O'Brien SJ. 1987b. Chromosome evolution of the Canidae. II. Divergence from the primitive carnivore karyotype. Cytogenet Cell Genet 44: 134-141.

Wienberg J, Stanyon R. 1998. Comparative chromosome painting of primate genomes. ILAR J 39:77 91.

Wijnen LMM, Grzeschik KH, Pearson PL, Meera Khan P. 1977. The human PGM-2 and its chromosome localization in man-mouse hybrids. Hum Genet 37:271-278.

Wipf L, Scackelford RM. 1942. Chromosomes of the red fox. Proc Natl Acad Sci U S A 28:265-268.

Wurster-Hill DH, Centerwall WR. 1982. The relationship of chromosome banding patterns in canids, mustelids, hyena, and felids. Cytogenet Cell Genet 34:178-192.

FIGURE 1 Gene map of the red fox (Vulpes vulpes) contains 35 biochemical loci. Gene symbol abbreviations are as follows: HPRT, hypoxanthine phosphoribosyl transferase; IDH1, isocitrate dehydrogenase-1; ITPA, inosine triphosphatase; LDHA, lactate dehydrogenase A; LDHB, lactate dehydrogenase B; ME1, malic enzyme-1; MDH1, malate dehydrogenase-1 (NAD dependent); MDH2, malate dehydrogenase-2 (NAD dependent); MPI, mannose phosphate isomerase; NOR, nucleolar organizer region; NP, purine nucleoside phosphorylase; OTC, ornithine carbamoyltransferase; PEPA, peptidase A; PGD, 6-phosphogluconate dehydrogenase; PGM 1, phosphoglucomutase- 1; PGP, phosphoglycolate phosphatase; PRNP, prion protein; PP, inorganic pyrophosphatase; SST, somatostatin peptide.

FIGURE 2 Syntenic genes and similar patterns of GTG banding on human, mink, cat, and fox chromosomes. The short arm of human chromosome 1 (HSA1p), the long arm of mink chromosome 2 (MVI2q), and the short arm of cat chromosome C1 (FCA C1p) exhibit similar banding patterns. Regional assignments of homologous genes are as follows: PGM1, PGD, ENO1 to HSA1p; the homologous mink genes to MVI2q; and the cat genes homologous to PGM1 and PGD to FCA C1. The human PGD and ENO1 genes are located very close to each other. In the fox, PGD was assigned to chromosome 2 (VVU2), but ENO1 and PGM1 were assigned to chromosome 12 (VVU12). The fox is the only known species to have these genes asyntenic. No region similar to Hsap1p, MVI2q, or FCA C1p was found on VVU2 or VVU12, nor was such a region found on MVI8 (containing the MDH 1, ACP1, ITPA, ADA genes), FCA A3 (the MDH1, ACP1, ITPA, ADA genes), HSA2p (containing the MDH1, ACP1 genes), or fox chromosomes VVU8 (containing the ACP1 gene), and VVU16 (containing the MDH1 gene). MVI8 and FCA A3 exhibit excellent banding pattern homology over the entire respective chromosomes. It is likely that they contain a region of homology with part of HSA2p. No region of similar banding was found on VVU16 or VVU8 (Rubtsov and others 1988).