Assessing the relative ages of admixture in the bovine hybrid zones of Africa and the Near

East using X-chromosome haplotype mosaicism.

Abigail R. Freeman*, Clive J. Hoggart§, O. Hanotte† and Daniel G. Bradley*

*Smurfit Institute of Genetics, Trinity College, Dublin 2, Ireland.

§Imperial College, London, United Kingdom.

†International Livestock Research Institute, PO Box 00100, Nairobi, Kenya.

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Genetics: Published Articles Ahead of Print, published on April 2, 2006 as 10.1534/genetics.105.053280

Running Title: Bovine X-chromosome haplotype mosaicism and linkage disequilibrium.

Keywords: admixture, haplotype mosaicism, linkage disequilibrium, Bos taurus.

Corresponding Author:

Professor Daniel Bradley

Smurfit Institute of Genetics,

Trinity College,

Dublin 2,

IRELAND.

Telephone: 00 353 1 608 1088

Fax: 00 353 1 679 8558

Email address: dbradley@tcd.ie

2

ABSTRACT

Historical hybridisation events between the two sub-species of cattle, Bos taurus and Bos

indicus, have occurred in several regions of the world, while other populations have remained

non-admixed. We typed closely linked X-chromosome microsatellites in cattle populations

with differing histories of admixture from Africa, Europe, the Near East and India.

Haplotype breakdown will occur as admixed populations age, and longer ancestral haplotypes

will remain intact in more recently admixed populations compared to older ones. We

genotyped male animals from these populations, obtaining unambiguous haplotypes, and

measured levels of linkage disequilibrium (LD) and ancestral mosaicism. Extensive LD,

likely to be the result of ongoing admixture, was discovered in hybrid cattle populations from

the perimeter of the tsetse zone in West Africa. A Bayesian method to assign microsatellite

allele ancestry was used to designate the likely origin of each chromosomal segment, and

assess the relative ages of admixture in the populations. A gradient of the age of admixture in

the African continent emerged, where older admixture has produced more fragmented

haplotypes in the south, and longer intact haplotypes, indicating more recent hybridisation,

feature in the northwest.

3

INTRODUCTION

Cattle have been domesticated at least twice from two distinct wild aurochs populations

(LOFTUS et al. 1994; BRADLEY et al. 1996; LOFTUS et al. 1999). The two extant taxa are Bos

taurus (taurine) and Bos indicus. The former were domesticated in the Middle East, Anatolia

and probably Africa about 10,000 years ago and are now found throughout northern Eurasia

and certain regions of Africa. Bos indicus or zebu cattle were domesticated from a different

variant of wild progenitor on the Indian sub-continent.

Substantial genetic exchange has taken place between the two types. Zebu cattle were

brought from southern Asia to the Middle East and the breeds that are found in this area

today are affected by hybridisation that took place perhaps over 3000 years ago (LOFTUS et

al. 1999). Nuclear genetic markers including Y-chromosome polymorphisms (BRADLEY et

al. 1994; HANOTTE et al. 2000) and autosomal microsatellites (MACHUGH et al. 1997) have

been used to show that there has been a widespread introgression of zebu cattle into the

African continent, which would have initially been inhabited only by African B. taurus, pure

examples of which remain only in certain parts of North and West Africa.

The genetic exchange between taurine and zebu cattle represents an appropriate model to

study the effects of post-admixture haplotypic decay through recombination, mutation and

genetic drift. Four of the domestic cattle hybrid zones; the Middle East, plus East, West and

Southern Africa, have breeds with roughly equal amounts, but different ages of admixture

(LOFTUS et al. 1999; HANOTTE et al. 2002). The first generation population created after an

admixture event will contain intact chromosomes from each of the inputting parental

populations. However, as mating occurs within the hybrid population these intact

chromosomes are mixed by recombination and gradually become more mosaic with each

4

generation. Therefore, the extent of genomic blocks undisturbed by hybrid recombination

will contain information about the history of admixture.

The examination of closely linked markers can be used to estimate how parental haplotypes

have been broken down over time in the hybrid population. Microsatellite allele frequency

spectra have been shown to differ sharply between B. indicus and B. taurus, facilitating the

assignment of ancestry at specific chromosome positions. Moreover extended haplotypes

may be unambiguously assigned to X-chromosomes in males.

Here we investigate differences in haplotypic mosaicism among 346 X-chromosomes typed

in male cattle from regions of differing phylogeographic history. We estimate microsatellite

allele ancestry at ten loci within a 10cM region to examine the breakdown of genomic blocks,

and assess levels of linkage disequilibrium (LD) in populations with differing histories of

admixture. We relate the different levels of haplotypic mosaicism and LD to the different

time depths and manners of introgression. For simplicity, we use the term LD to describe

both traditional linkage disequilibrium, and pairwise disequilibrium between non-linked

markers.

5

MATERIALS AND METHODS

346 male cattle DNA samples from 22 breeds were obtained as described previously (Table

1) (MACHUGH et al. 1997; LOFTUS et al. 1999; RITZ et al. 2000; HANOTTE et al. 2002). The

approximate locations of the sample sites and previously calculated admixture estimates (mR)

are shown in Figure 1 (MACHUGH et al. 1997; LOFTUS et al. 1999; HANOTTE et al. 2002;

FREEMAN et al. 2004). For LD analysis, the novel data obtained here were combined with

microsatellite data from previously published studies (MACHUGH et al. 1997; LOFTUS et al.

1999; HANOTTE et al. 2002; FREEMAN et al. 2004). Of the 22 breeds, two were unadmixed

European B. taurus, three were primarily unadmixed West African B. taurus and two were

primarily unadmixed B. indicus. The remaining 15 breeds were known to have been formed

by admixture between Bos taurus and Bos indicus. Figure 1 shows the geographic location of

the breeds

10 X-chromosome microsatellites were selected from the Meat Animal Research Center

(MARC) database http://sol.marc.usda.gov/genome/genome.html (BISHOP et al. 1994; SUN et

al. 1994; KAPPES et al. 1997). All of the microsatellites selected have been linkage mapped

to a 10 cM region on the X chromosome (KAPPES et al. 1997; SØNSTEGARD et al. 1997;

SØNSTEGARD et al. 2001; IHARA et al. 2004). Six of the markers have also been included in

radiation hybrid maps of the bovine X chromosome (BtaX) (AMARAL et al. 2002; WILLIAMS

et al. 2002), and all but one of the markers can be found in the Pre! ensembl bovine genome

assembly (http://pre.ensembl.org/Bos_taurus/index.html). Approximate distances between

markers and marker order were taken from the genome assembly after observing agreement

between this and the linkage and radiation hybrid maps for most markers. To control for

potential mis-orientation of contigs in the preliminary assembly, a comparison with the

syntenic chromosomal region in humans was examined. Again, order seemed to be mostly

6

conserved. Thus, for the purposes of this paper, the marker order and positions listed in

Table 2 were used. Two methods of genotyping were used, with common standards enabling

combination of the data. Populations marked with an asterisk in Table 1 were genotyped

using a radioactive PCR method described in MacHugh et al (1997), while the remaining

populations were genotyped using an ABI 377™ system following the protocol of Hanotte

and colleagues (2002).

Pairwise linkage disequilibrium between locus pairs was carried out using an extension of

Fisher’s exact test implemented by the GENEPOP program (v3.2a) (RAYMOND and ROUSSET

1995). Populations were defined according to the regional groups in table 1. For a given pair

of loci within one population, the possible genotypes are represented by GENEPOP in a

contingency table. For each locus pair within each population, the unbiased estimate of the

p-value is calculated using a Markov chain as the sum of the probabilities of all tables (with

the same marginal values as the observed one) with a lower or equal probability than the

observed table.

Statistical inference on admixture and the ancestry crossover rate in the admixed cattle

populations was made in the computer program ADMIXMAP by simulating posterior

samples of the model parameters using Markov chain Monte Carlo simulation (HOGGART et

al. 2003; HOGGART et al. 2004). ADMIXMAP implements a full Bayesian hierarchical

model for admixture at the population, individual and locus level and also models the allele

frequencies in the unadmixed subpopulations that have contributed to the admixed population

under study. The stochastic variation of ancestry between loci on each chromosome is

modelled by independent Poisson arrival processes, one for each unadmixed population

contributing to the admixed population under study, in our case B. taurus and B. indicus.

7

Thus inference is made on parameters representing population and individual admixture,

indicator parameters for locus ancestry for all loci and all individuals, parameters of

multinomial distributions representing allele and haplotype frequencies in the two

populations and τ, the sum-of-intensities of the two Poisson arrival processes. Whilst

admixture was modelled separately for each individual in a population (independent and

identically distributed from the population level admixture distribution) τ was assumed to be

the same for all individuals in the population.

For an individual with admixture proportion θ from B. taurus the intensities per Morgan of

the arrival processes for B. taurus and B. indicus are θτ and τθ)1( − respectively. The

ancestry crossover rate ρ (the rate per Morgan of transitions between ancestry of one

subpopulation to ancestry of the other) is given by )2 1( θτθρ −= , thus the larger τ the more

frequent the crossovers. Since the X chromosome only undergoes recombination in female

gametes, a different τ will be applicable to the X chromosome than the autosomes. However,

in our data sets the autosomal markers are unlinked thus τ was estimated from the linked

markers typed in the 10cM region of the X chromosome alone. In contrast individual

admixture proportions (θ) were estimated from both the autosomal and X chromosome

markers. When introgression occurs in a single pulse the expectation of τ on the autosomes is

equal to the number of generations since admixture, (FALUSH et al. 2003). However,

admixture in the cattle populations is not believed to have occurred as a single event but

rather through continuous gene flow of B. indicus into B. taurus populations. The effect of

continuous gene flow on τ is unclear thus it cannot be interpreted as the number of

generations since admixture but as a relative measure of the age of admixture among the

populations studied. Given the lack of interpretation of τ it was assigned an uninformative

prior, log τ uniform. Prior distributions for population specific allele frequencies for Bos

8

taurus and Bos indicus were set from allele counts in the five Bos taurus populations (both

European and west African) and two Bos indicus populations listed in Table 1. Whilst it is

known that the West African Bos taurus populations contain a degree of indicus admixture

this is very small (mR = 3%, 7% and 10% in Guinean N’Dama, Guinea Bissau N’Dama and

Somba respectively) and thus will have minimal effect on the allele counts. Furthermore,

specifying prior distributions from the allele counts rather than fixing them at their mean

values allows for error in the frequencies to be accounted for. Uninformative uniform priors

were assigned for the population level admixture.

The ADMIXMAP analyses assumes that all LD between markers can be attributed to

admixture LD and LD due to ancestral mosaicism but ignores background LD. The X

chromosome markers used in our analyses are between 0.1 Mb and 2.5 Mb apart (Table 2),

and at such distances it is a reasonable approximation to ignore background LD.

Statistical testing was carried out using SPSS (v. 12.0) and values of τ were plotted with

sampling location using the ARCVIEW GIS software package. Equal interval contours join

areas with similar values of τ (Environmental Systems Research Institute, ESRI, Redlands,

CA). A synthetic map was finalised using the Adobe Illustrator (v. 8.0) package.

9

RESULTS

A set of ten linked X-chromosome microsatellites that have been mapped within a 10cM

region of BtaX were genotyped in 346 male cattle samples from geographical locations

shown in Figure 1 and detailed in Table 1. The data were combined with unlinked autosomal

microsatellite genotypic data from previous publications.

The strength of pairwise linkage disequilibrium (LD) between all typed markers was

estimated and the results divided into two groups according to the significance of association

between pairs of markers: p<0.05 and p<0.01. Figure 2 shows matrices of LD significance

levels for all possible pairwise comparisons of the 10 X-chromosome loci in their

chromosomal order, and for the unlinked markers. The samples were divided into regional

groupings according to Table 1, excluding the Zebu Malagasy breed which does not easily

fall into an appropriate category and which is only represented by 4 individuals. The

unlinked autosomal microsatellite markers provide a measure of the background levels of LD

in each regional group. LD measures for linked markers are shown on the left hand side of

the heavy black line, in the order that they are located on the X-chromosome.

In general, and as expected, physically linked markers tend to have higher levels of

association than markers from different chromosomes. This trend is evident in all groups

except European B. taurus. This group exhibits slightly lower levels of LD among linked

markers than unlinked markers, but the difference is small. The trend is apparent in all of the

hybrid populations (Near Eastern, West African, East African, South African), but most

especially in the West African hybrid population which exhibits especially high levels of LD

among the X-chromosome microsatellites. Nearly 45% of all possible pairwise comparisons

10

between linked markers are in LD at a significance level of 0.01 in this group. This is over

three times as many as is seen in any of the other population groups.

Estimates of τ obtained from ADMIXMAP enable comparison of time since admixture

among the different sampled populations, but cannot be taken as absolute values, as a single

event is unlikely to be an accurate model of past hybridisation processes. Values of τ are

shown on a synthetic contour map (Figure 3). Despite the relatively small number of hybrid

breeds (fifteen), some geographical congruency can be observed for values of τ.

The posterior means and standard deviations of the sum-of-intensities parameter τ for the

admixed breeds are shown in Table 1. We see that within the African continent there are two

main trends among the hybrid populations. Firstly, values of τ tend to be higher in the south

of the continent, and secondly there is a decreasing cline in values of τ from east to west in

the northern part of the continent. To test for significant difference between hybrid breeds in

the north and south we pooled the estimates of τ using the inverse variance method (DEEKS et

al. 2001). The pooled estimate of τ for hybrid breeds in the northern part of the continent

(Mbororo, Gobra, Sokoto Gudali, Arashie, Kuri) is 7.6 (standard deviation 1.66), and the

pooled estimate for hybrids in the south (Ogaden, Ankole, Afrikaner, Nguni, Watusi, Zebu

Malagasey, Barotse, Kavango) is 13.9 (standard deviation 1.85). The test for difference

between the means of the groups is significant, p=0.012. The pooled estimate for τ in the two

Near Eastern hybrids is 15.4 (standard deviation 3.54), with the highest value being observed

in the eastern-most Iraqi breed (τ =17.8).

11

DISCUSSION

Cattle populations represent an ideal model for the study of admixture generated linkage

disequilibrium and haplotype decay, as it is well established that they were domesticated

from divergent wild progenitors leading to two main interfertile lineages: Bos taurus and Bos

indicus. Since domestication, the two types have been combined in many different hybrid

breeds but there are also pure extant taurine and zebu populations that provide good proxies

for the original ancestral populations (MACHUGH et al. 1997).

The amount of admixture in the breeds used in this study had previously been estimated using

autosomal microsatellites (MACHUGH et al. 1997; LOFTUS et al. 1999; HANOTTE et al. 2002;

FREEMAN et al. 2004)(Figure 1). Here we set out to examine the pattern of haplotype decay

in the bovine genome after admixture by genotyping closely linked microsatellite markers.

The level of divergence between the contributing parental populations will determine whether

it is possible to assign definite ancestry to an allele in a hybrid individual. In a previous

study, we found that over 80% of microsatellite alleles surveyed at 20 loci in European, Near

Eastern and Indian cattle populations showed greater than 50% frequency differential (scaled

by the absolute allele frequency) between B. indicus and B. taurus (KUMAR et al. 2003). All

of the markers used here are located within a 10cM region of the X-chromosome and were

typed only in males. This allowed the unambiguous assignment of a haplotype to an

individual without pedigree information.

We found LD in this region of the X-chromosome in all of the cattle populations, and

between linked and unlinked markers in all of the populations except for the Near Eastern

hybrids. These results concur with Farnir et al. 2000 who surveyed 284 autosomal markers

and found that intra-chromosomal LD extends over several tens of centiMorgans and is

12

common between non-syntenic loci in dairy cattle populations. In that case, the high levels

of overall LD may have been a result of a small effective population size, (LONJOU et al.

1999) which could be as low as 50 for 1.2 million Holstein-Friesian females, or admixture

(FARNIR et al. 2000).

Recurrent introgression has been cited previously as the cause of higher than expected levels

of LD in African American human populations (LAUTENBERGER et al. 2000). Here, the very

high level in the West African hybrid population may partially be explained by recent and

continuous gene flow to this region (PFAFF et al. 2001). Pfaff and colleagues argue that

maintenance of LD over relatively large (~10 cM) chromosomal segments is a not typical of

a single admixture event, but is a characteristic of a continuous gene flow pattern of

admixture. While the initial introgression of B. indicus chromosomes into Africa is likely

ancient, zebu-taurine admixture in West Africa probably began only recently in many

populations where it had previously been constrained by the tsetse/trypanosomiasis

infestation zone; a barrier to introgression. Understanding the patterns of LD in the hybrid

populations from the surround of the West African tsetse zone will prove useful if these

populations are used to map genes which influence disease traits (LAAN and PAABO 1997).

In the absence of mutation, every portion of a chromosome in a hybrid individual should be

directly traceable to one of the individual’s ancestors. When two distinct gene pools meet, a

mosaic genome is produced (CHAPMAN and THOMPSON 2002), containing intact segments

from each contributing founder populations (NORDBORG and TAVARE 2002). Genomes that

have undergone recent hybridisation will have largely intact ancestral haplotypes, while

recombination will act to disrupt these haplotypes in subsequent generations (WIEHE et al.

2000). If admixture is ancient, reciprocal recombination, genetic drift and mutation will have

13

eroded the association between markers from parental populations and a greater density of

markers will be required to detect significant associations.

Here we employed a Bayesian method to assign probable ancestry to each allele in every X-

chromosome haplotype. The amount of fragmentation in haplotypes was assessed by the

posterior distribution of τ, and this was used as a measure for the age of admixture. The mean

values of τ (Table 1) for each of the cattle breeds are between 4.27 and 21.3 which is

substantially less the number of generations since admixture is believed to have first

occurred. However, as discussed earlier, in the case of cattle a single admixture event may

not accurately model the historical reality and τ only has expectation equal to the number of

generations since admixture in the case of a single admixture event. Furthermore, the prior

has the effect of favouring small values of τ.

The values obtained appear to show some geographical consistency, whereby most hybrid

populations from southern Africa and the Near East appear to have experienced older

admixture than those in the more northern part of the African continent. Furthermore there is

a decline in age of admixture from east to west in the latter. The differences between the east

and the west of the continent support a scenario of earlier introgression of B. indicus material

into the former populations, as the degree of recombination would be expected to accumulate

with time. This finding is supported by high levels of LD in West African hybrids, and

previous work which has shown that the major B. indicus introductions occurred in the East:

probably in the region of the Horn (HANOTTE et al. 2002).

The difference in τ values between the south and the north of Africa may be a signal of

different histories of cattle in the two regions. However, it is well established that the earliest

14

Our sample from Madagascar suggests that admixture here was older but this sample is very

small, n=4, and should not be over-interpreted. There is limited archaeological evidence for

early arrival of B. indicus cattle to Madagascar and zebu cattle occupy a place of prominence

Based on archaeological evidence the origins of B. indicus introgression in Africa are

controversial. Some authors have suggested that they were found as early as 3500-2000 BP

(HASSAN 2000; MARSHALL 2000; PARIS 2000), but these studies are inconclusive, and the

earliest verified archaeological date for their presence is in the Horn of Africa in the second

century AD (MARSHALL 2000). The archaeological record shows that B. taurus cattle were

present in northern Africa and the Sahara from approximately 6000 BP (HASSAN 2000), and

spread as far south as Kenya by 3000 BP (HASSAN 2000; MITCHELL 2005). Further

southward movement may have been hampered by ecological factors such as the many sub-

Saharan disease challenges to which domestic cattle would be maladapted (GIFFORD-

GONZALEZ 2000; MITCHELL 2005), and relatively few cattle were present in the south until

1000 AD (SMITH 2000). It is therefore possible that nomadic pastoralists in eastern Africa

were herding animals comprised of the two cattle lineages before the expansion of domestic

cattle to the south.

domesticated cattle of Africa were found in the north of the continent and were B. taurus in

nature, and that introgression of B. indicus into the continent via the Horn of Africa was

secondary. Our results suggest that these earliest hybrids may have been the animals that

later gave rise to southern populations. This scenario implies that B. indicus introgression

occurred before pastoralism spread extensively southwards and indeed, despite extensive

sampling, there is no evidence for the presence of pure African B. taurus in the south of the

continent (HANOTTE et al. 2002).

15

16

in Malagasy ritual and culture, which is compatible with an extensive history on the island

(HEMMER 1990; VAN DER ZWAN and EVERS 1998).

The employment of haplotypic mosaicism as an indicator of the antiquity of admixture

receives some confirmation from the Near Eastern results. The region between the primary

centres of B. indicus and B. taurus domestication is an area of ancient interaction between the

two lineages. As such, the sample from Iraq yields one of the highest values of τ within the

haplotypes.

Acknowledgements

This material is based on works supported by Science Foundation Ireland under grant no. 02-

IN.1-B256. AR Freeman was funded by a Wellcome Studentship in Biodiversity number

054275/Z/98/Z. We would like to thank David Lynn, Colm O’hUigin, Paul McKeigue and

Mark Shriver for helpful discussions and Claude Gaillard for provision of samples.

17

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Table 1 Cattle breeds included in the present study. N is the number of individuals in each

sample, E(τ) and SD(τ) are the posterior mean and standard deviation of τ respectively and τ

is a measure of the age of admixture. The asterisks mark those breeds that were genotyped

using radioactively labelled PCR products.

Breed name N E(τ ) SD(τ) Regional designation Aberdeen Angus* 29 n/a n/a European Bos taurus

Charolais* 33 n/a n/a European Bos taurus

N’Dama (Guinea)* 36 n/a n/a West African Bos taurus

N’Dama (Guinea Bissau)* 22 n/a n/a West African Bos taurus

Somba* 10 n/a n/a West African Bos taurus

Gobra* 32 6.67 2.70 West African hybrids

Sokoto Gudali* 13 9.90 4.62 West African hybrids

Mbororo* 9 4.27 3.24 West African hybrids

Kuri* 13 13.00 5.01 West African hybrids

Arashie 16 10.71 4.79 East African hybrids

Ogaden 18 10.23 4.76 East African hybrids

Ankole 14 7.99 4.92 East African hybrids

Watusi 3 15.45 7.23 East African hybrids

Barotse 16 18.90 5.04 Southern African hybrids

Kavango 12 21.30 4.78 Southern African hybrids

Nguni 9 15.27 5.87 Southern African hybrids

Afrikaner 14 8.87 4.28 Southern African hybrids

Zebu Malagasy 4 17.92 6.65 Madagascar

Anatolian Black* 12 13.13 5.05 Near Eastern hybrids

Iraqi* 16 17.76 4.98 Near Eastern hybrids

Nellore* 12 n/a n/a Bos indicus, of Indian descent

Ongole* 3 n/a n/a Indian Bos indicus

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Table 2. Primer sequences and Optimal PCR conditions for ten X chromosome microsatellites. Magnesium Chloride concentration is shown

by [MgCl2] and annealing temperature is denoted by TA

Locus name

Approximate chromosome position (Mb)

Accession number UniSTS Primers (5’-3’) Amplicon

size PCR

[MgCl2] TA PCR

BMS1616 7 G18677 27189 CAGTGTGTATAGCATGATTCCG AGTTGGTCTGCATTCATACATTAA 92-116 1.25 54-55ºC

BL1045

9.2 251338 TGCCAGAACAAGTCACAAGC CTCCAAGGGTGTCTCTATCCC 92-100 1.5 59ºC

UWCA19 9.4 U01794 251002 CTGTGATAACCTATATGGGAAAGGA GAGGACAGCAAAGTGATTCAGT 97-105 1.5 58ºC

BL1098 10 251324 CCACAACTTCCAGAAGCCTC CAGAAACCACCCAAACTAACC 203-235 1.25 56ºC

BMS960 11 G18760 74452 TGTAAATAGCTCCCCTCCTGC CTAGCAATTTGTTGTTCATGGC 106-128 1.75 57ºC

XBM701 11.7 251432 TTCCCCTTGAAATCATCTGG TCCAAGTTACCAAAATTGACCC 148-168 2.0 58ºC

BMS2713 12.2 G18462 69721 TGAATACCTGTTTCCAGCCC CTCCTAAGTCCAGGAAGCCC 143-155 1.5 56ºC

XBM7 251010 CTGTATTAGAGTTCCCTGGAGAAA GCCAACATGCCCTGTAGAAT 180-202 1.5 57ºC

XBM361 14.8 251341 ACACACTGAGAATTAAACTACAAAAACCACACAACTGAAGCCACTTTAAGC 150-162 1.25 58-59ºC

BMS649 16.3 G18728 37262 CTCGGACTCATAACGCACAT CGAGCAACAAGAGTGAAGGT 116-134 1.75 59ºC

23

FIGURE LEGENDS

Figure 1. Map showing approximate sampling sites for cattle populations included in the current

study. The areas of the pie charts are proportional to the number of animals in the population.

The pie charts represent a previously calculated mR value for each breed which is a measure of

Bos taurus/Bos indicus admixture which has been calculated using European B. taurus and

Indian B. indicus breeds as parental populations. The Nellore breed was sampled in Brazil, but is

comprised of Ongole individuals that were brought to South America (FELIUS 1995; RITZ et al.

2000).

Figure 2. Linkage disequilibrium plots showing LD assessed by the significance of allelic

associations at pairs of loci. The lightest grey shaded areas represent untyped markers. Grey

and black squares mark locus pairs with alleles associating with p-values of <0.01 and <0.05

respectively. The matrix plots facilitate comparison of LD levels among the different regional

groups. Additionally, the differences between LD among the sets of linked markers and

unlinked markers are also evident. To further clarify the differences both between regional

groups and between sets of linked and unlinked markers, the percentages of significant

associations (at the two designated levels) are also shown in two histograms. The cattle

populations are grouped according to Table 1.

Figure 3. Map showing values of τ. τ is the number of generations since a single admixture

event admixture. In these cattle populations admixture probably occurred in a continuous

manner, therefore τ is interpreted as a relative measure of the age of admixture among the

populations studied (see Materials and Methods). Higher values of τ are shaded with darker

shades of grey and suggest older admixture in those areas.