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Arch. Tierz., Dummerstorf 50 (2007) 1, 07-24
Institute for Animal Breeding and Genetics, University of Veterinary Medicine Hannover, Germany
DOROTHEE STÜBS and OTTMAR DISTL
Mapping the horse genome and its impact on equine genomics for
identification of genes for monogenic and complex traits – a review
Abstract
Since the beginning of investigation in the horse genome in the early nineties, there has been a great progress,
especially during the last five years. At the beginning the exploration of monogenic hereditary diseases was one
of the main aims, and the causal mutations of several diseases in the horse have been unravelled. The inheritance
of coat colours has been explored very detailed, and there exist gene tests for different coat colours. Information
about coat colours and inherited diseases is very important for the breeders and helps avoiding the appearance of
lethal genetic factors or undesirable diseases. The most important achievements of horse genome analysis were
well-developed linkage, radiation hybrid and cytogenetic genome maps including more than 2950 loci. These
maps support comparative analysis of equine hereditary diseases. The present known gene mutations for five
diseases in horses have human homologs. Studies on multifactorial diseases such as osteochondrosis and
navicular bone disease and on fertility and temperament are underway. At the moment, the whole equine genome
is sequenced as it has been done for the human genome and also for other animal genomes. Horse breeding will
greatly benefit from identification of QTL for multifactorial traits and gene mutations for congenital anomalies,
diseases and performance traits.
Key Words : horse, genome, equine maps, coat colours, inherited diseases
Zusammenfassung
Titel der Arbeit: Kartierung des Pferdegenoms und funktionelle Genomanalyse zur Identifizierung
monogenischer und komplexer Merkmale – ein Überblick
Seit den ersten Arbeiten zur Aufklärung von Genen beim Pferd in den 90er Jahren wurden große Fortschritte in
der Genomanalyse beim Pferd gemacht. Für fünf monogen vererbte Krankheiten konnten die verantwortlichen
Genmutationen aufgeklärt werden. Die Vererbung der Fellfarben auf populationsgenetischer Ebene ist beim
Pferd weitgehend geklärt. Für einige Fellfarben, wie die Fuchsfarbe, den Sabino, die Tobiano-Scheckung und
den Aufhellungsfaktor (Cream Allel) am Albino Genort, konnten Gentests entwickelt werden. Diese
Erkenntnisse über die Farballele sind für die Pferdezucht wichtig, und Gentests für die Erkennung von
Anlageträgern helfen, das Auftreten von Erbfehlern und Krankheiten zu vermeiden. Die bisher wichtigsten
Fortschritte in der Genomanalyse waren die Entwicklung von genetischen, zytogenetischen und Radiation
Hybrid Karten, damit Genorte für monogen und multifaktoriell vererbte Merkmale (QTL) kartiert und
anschließend vergleichend analysiert werden konnten. Bisher wurden mehr als 2950 Genloci auf diesen Karten
kartiert. Arbeiten zu multifaktoriellen Krankheiten wie Osteochondrose und Podotrochlose sowie zur
Fruchtbarkeit sowie zu dem Temperament sind auf dem Weg. Momentan wird das Pferdegenom komplett
sequenziert wie dies bereits für den Menschen und andere Haustiere durchgeführt wurde. Die Pferdezucht wird
davon für die Identifizierung von QTL und Aufklärung von Genmutationen für angeborene Anomalien,
Krankheiten und Leistungsmerkmale erheblich profitieren.
Schlüsselwörter : Pferd, Genom, Genkarten, Fellfarben, erbliche Krankheiten
Introduction
The first animal genome sequence had been completed already when scientists
decoded the sequence of the nematode C. elegans (BETHESDA, 1998). The
exploration of mammalian genomes, especially in farm animals, proceeded very fast,
and there are several genomes of mammalian animals completed today (FADIEL et al.,
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STÜBS; DISTL: Horse genome
2005). Genome sequencing in other domestic animals, particularly in pets, forged
ahead in the last few years, too, because it was a matter of particular interest for
breeders. The chicken, dog, and cow genome have been wholly sequenced and work
has started in the pig and horse (BETHESDA, 2004).
Having the information on the complete genome sequence is the key on understanding
physiological and pathological processes and may revolutionize health research. While
scientific investigations had to be confined mainly to the analysis of phenotypic data
and pedigrees for familial diseases, today techniques include fluorescence in situ
hybridisation (JOHN et al., 1969), somatic cell hybrid panels and radiation hybrid
panels (STRACHAN and READ, 1996), comparative mapping (O`BRIEN and
GRAVES, 1991), sequence and mutation analysis, gene expression using microarrays
or quantitative real time techniques.
New information about mapped genes or markers is shortly posted on one of the
databases in the internet and is free available (HU et al., 2001).
The progress in investigation of the horse genome especially in the last ten years is the
result of concentrated and well-organized scientific work all over the world: 1995 was
the year of the foundation of the International Equine Gene Mapping Workshop. Since
1997 this foundation works together with the Horse Technical committee of the
National Research Sponsored Project-8 (NRSP-8) of the United States Department of
Agriculture National Animal Genome Research Program (USDA-NAGRP) with the
aim to decipher the horse genome as soon as possible.
Organization of the horse genome
The diploid genome of the domestic horse ( Equus caballus ) consists of 64
chromosomes, and 13 pairs of the autosomes are biarmed, while the other 18 pairs of
the autosomes are acrocentric. The X chromosome is large and metacentric, the Y
chromosome small and submetacentric. It is remarkable that the equus przewalski
genome consists of 66 chromosomes and that 12 pairs of its autosomes are metacentric
and 20 pairs are acrocentric. One pair of the horses` acrocentric chromosomes (ECA5)
contains two of the przewalski`s chromosomes (EPR23 and EPR24), hence it is
possible to mate przewalski and domestic horse and to produce fertile offsprings
(AHRENS and STRANZINGER, 2005).
Cytogenetic map
Physical identification of loci of interest can be achieved by fluorescence in situ
hybridisation (FISH). The first fluorescence in situ hybridisation in the horse was
reported by OAKENFULL et al. (1993) with the mapping of the alpha-globin gene
complex to horse chromosome (ECA) 13. BREEN et al. (1997) localized the first
equine markers when they mapped 18 microsatellites. RAUDSEPP et al. (1997)
cytogenetically mapped genes to the genome of the donkey. LEAR et al. (2001),
RAUDSEPP et al. (2002), GODARD et al. (2000) used goat BAC clones to localize
44 coding sequences on the equine FISH map. LINDGREN et al. (2001) localized
another 13 genes by FISH and somatic cell hybrids. One of the first most extensive
FISH-map included 136 genes (MILENKOVIC et al., 2002), and to this cytogenetic
map further 165 genes were added by FISH and/or radiation hybrid (RH) mapping
(PERROCHEAU et al., 2005). The equine gene map now contains 713 genes, 441 on
the RH map, 511 on the cytogenetic and 238 on both maps.
Arch. Tierz. 50 (2007) 1
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Several of gene groups attracted special interest for mapping in different animals, such
as immunity-related genes: GUSTAFSON et al. (2003) constructed an ordered BAC
contig map of the equine major histocompatibility complex (MHC) and
TALLMADGE et al. (2003) mapped the ß2-microglobulin gene that is responsible for
the light chain of the MHC to ECA1q23-q25. MUSILOVA et al. (2005) mapped
another 19 horse immunity-related loci by FISH. NERGADZE et al. (2006) localized
the IL8 gene on ECA3q14. Others mapped genes related to the bone-, cartilage- and
collagene-metabolism. Genes eventually responsible for bone- or collagene-related
hereditary diseases in the horse were mapped by HILLYER et al. (2005), MÜLLER et
al. (2005), BÖNEKER et al. (2005) and DIERKS et al. (2006).
Synteny and radiation hybrid maps
Synteny mapping and radiation hybrid mapping in the horse work analogously to those
in humans, but the utilization of these techniques began more than 20 years later than
in humans. While there had been five synteny panels in the horse since 1992 (LEAR et
al., 1992; WILLIAMS et al., 1993; BAILEY et al., 1995; RANEY et al., 1998), the
most important one was the UC Davis panel generated by SHIUE et al. (1999). It
included 450 markers and represents the origin for the developed horse-human
comparative maps (CAETANO et al., 1999).
A 3000-rad panel was developed by KIGUWA et al. (2000). Preliminary comparative
maps of horse chromosomes 1 to10 were created using this panel. A great progress
was the more extensive 5000-rad whole-genome-radiation hybrid panel that was
constructed using 92 horse x hamster hybrid cell lines and 730 equine markers (type I
and type II markers) (CHOWDHARY et al., 2003). This panel was one of the most
essential tools to construct high-resolution and comparative maps for ECA11
(CHOWDHARY et al., 2003), ECAX (RAUDSEPP et al., 2004), ECA17 (LEE et al.,
2004), ECA22 (GUSTAFSON-SEABURY et al., 2005), ECA4 (DIERKS et al., 2006),
equine homologs of HSA19 on ECA17, ECA10 and ECA21 (BRINKMEYER-
LANGFORD et al., 2005) and equine homologs of HSA2 on ECA6p, ECA15 and
ECA18 (WAGNER et al., 2006).
Comparative map
Comparative maps between horse and human and horse and mouse are the most
important ones. There also exist comparative maps between horse and other
mammalians e.g., between horse and donkey (RAUDSEPP et al., 1997, 1999, 2001;
RAUDSEPP and CHOWDHARY, 2001), horse and mountain zebra (RAUDSEPP et
al., 2002 ) and E. caballus and E. przewalski (MYKA et al., 2003). Comparative
chromosomal painting (Zoo-FISH) helps identifying conserved chromosomal
segments among species. Reciprocal painting of chromosomes and segments was
performed to compare the horse genome with that of the donkey (RAUDSEPP et al.,
1996; CHOWDHARY et al., 1998; YANG et al., 2004 ) .
One approach to localize genes and to expand the comparative gene map between
horse and man is to pick genes or DNA-/RNA-sequences from genes physically
mapped and cloned in humans for screening filters of a genomic horse BAC library.
After verification of the selected gene sequence on the genomic equine BAC clone,
FISH is used for mapping this BAC clone on horse metaphase spreads. There are three
equine genomic BAC-libraries available: the equine BAC library of the Children`s
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STÜBS; DISTL: Horse genome
Hospital of Oakland Research Institute, CHORI-241 (GODARD et al., 1998), one
generated by the Texas A&M University (TAMU) and the other by the Institut de la
National Recherche Agronomique (INRA-LGBC library) (CHOWDHARY et al.,
2003).
CAETANO et al. (1999) published an extensive comparative map between horse and
man containing 68 equine type I loci. The most topical comparative map between
horse, donkey and human was generated by cross-species painting (YANG et al.,
2004). The human-donkey map was the first genome-wide comparative map between
these species generated by chromosome painting. This horse-human comparative map
agrees basically with the comparative map of CHOWDHARY et al. (2003), except
from varying results especially for ECA1. For several horse chromosomes, detailed
comparative maps have been developed. A detailed physical map of the equine Y-
chromosome was generated (RAUDSEPP et al., 2004) and compared to the human and
other mammalian species with the result that the equine Y-chromosome shows the
most homology with the Y-chromosome of the pig. A comparative RH map for
ECA17 was developed by LEE et al. (2004), and there exists a detailed RH map of
ECA22 comparing the equine loci to human, dog, cat, bovids, elephant and bat,
rodents, dolphins and other mammalians (GUSTAFSON-SEABURY et al., 2005).
This map covered estimated 64 Mb of physical length of chromosome 22 (831 cR) and
the 83 markers had an average distance of 10 cR. It was remarkable, that the most
common configuration for the ECA22 homologs consisted for Hartmann’s zebra as
expected, but also for carnivores, hippos, primates, rabbits, squirrels and bats. The
conserved synteny between the compared animals can be taken as an indicator of the
degree of putative ancestral relationship between the different species.
BRINKMEYER-LANGFORD et al. (2005) compared equine homologs of HSA19 to
other mammals using 89 loci for the 5000-rad-RH-panel and four loci for FISH. The
homologs to HSA19 were localized on ECA7, 21 and 10. This comparison
demonstrated an overview of the evolution of putative ancestral HSA19 chromosomal
segments in more than 50 mammalian species.
PERROCHEAU et al. (2006) developed a medium density horse gene map using 323 genes
evenly distributed over the human genome. They mapped 87 genes by FISH and 186 genes by
the equine 5000-rad RH panel and thereby detected previously unknown homology between
ECA27 and HSA8 as well as between ECA12p and HSA11p.
Linkage map
Genetic and physical assignment of equine microsatellites began almost ten years ago.
BREEN et al. (1997) isolated 20 equine microsatellites from a genomic phage library
and mapped them by linkage mapping and FISH. GODARD et al. (1997) assigned 36
new horse microsatellites, 11 of them from plasmid libraries and 25 from a cosmid
library.
The first equine linkage map included 140 markers (LINDGREN et al., 1998) and a
more extensive linkage map consisted of 161 loci in 29 linkage groups for 26
autosomes (GUERIN et al., 1999). The first map covering all autosomes and the sex-
chromosomes using 353 microsatellite-markers was generated by SWINBURNE et al.
(2000). The microsatellites were mapped to 42 linkage groups and covered 1780 cM of
the horse genome.
A second generation linkage map was published by the International Equine Gene
Mapping Workshop (GUERIN et al. 2003). This linkage map was based on testing
Arch. Tierz. 50 (2007) 1
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503 half-sibling offspring from 13 sire families. The map included 344 markers in 34
linkage groups representing all 31 autosomes, but neither the X- nor the Y-
chromosome. The linkage groups covered 2262 cM with an average interval between
loci of 10.1 cM, ranging from 0 to 38.4. Another 61 new horse microsatellite loci were
assigned by SWINBURNE et al. (2003). TOZAKI et al. (2004) reported the
characterization of 341 newly isolated microsatellite markers of which 256 were
assigned to equine chromosomes.
PENEDO et al. (2005) added 359 microsatellites to the second generation linkage
map. Alltogether, this linkage map consisted of 766 markers assigned to the 30
autosomes and the X-chromosome spanning 3740 cM with an average marker density
of 6.3 cM. SWINBURNE et al. (2006) generated a linkage map including 742 markers
in 32 linkage groups according to the 31 autosomes and the X-chromosome. All
linkage groups together span 2772 cM with an average interval of 3.7 cM between the
markers.
Chromosomal abnormalities
There exist several chromosomal abnormalities in the horse. Of great interest and
therefore well analyzed are those concerning the sex-chromosomes and influencing the
fertility of mares:
The most common abnormality in mares, X0 Gonadal Dysgenesis (X monosomy), is
characterised by the lack of one X-chromosome (63,X0) and was primarily described
by HUGHES et al. (1975). This defect was accompanied with infertility of the affected
mare. POWER (1986) reported another abnormality in horses, the XY Gonadal
Dysgenesis. The affected horses` phenotype is female, but their genotype is 64,XY
leading to infertility (BOWLING et al., 1987). There exist also infertile mares with the
genotype 65,XXX (CHANDLEY et al., 1975). KAKOI et al. (2005) published a
statistical evaluation of sex chromosome abnormalities and analyzed data of 17471
light-breed foals with the result that 0.15 % of the analyzed population showed an
XO-, 0.02% an XXY- (and/ or mosaics/chimaeras) and 0.01% an XXX-genotype.
Unlike the sex chromosomes, chromosomal abnormalities of the autosomes are rarely
encountered. Cases of trisomy are similar to human Downs and associated with mental
retardation failure to thrive (LEAR et al., 1999).
Coat colours
There exist several genetically characterized major genes that influence the coat colour
of a horse: Agouti-gene, Extension-gene, Cream dilution gene, Tobiano-gene, White-
gene, Greying hair-gene, Dun-gene, Roan-gene, Sabino-gene, Silver-gene, and the
Leopard Complex-gene. Knowledge about the inheritance of coat colours is very
important for the breeders, so it is comparatively well explorated and for several genes
exist gene tests, too. Chromosomal locations and mutations of genes associated with
equine coat colours are summarized in Table.
The White-gene and the Greying hair-gene can mask all the other coat colour genes, so
they are mentioned first.
White-gene (W-gene) and Sabino spotting
Horses with the dominant allele “W” lack pigment in skin and hair, so their coat colour
is white at birth, but eyes are dark or sometimes blue. The W-mutation was mapped to

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