+Supplementary Materials

Genetic Structure of Omani Goats Reveals Admixture among Populations from

Geographically Proximal Sites

Nasser Ali Al-Araimia*, Raed Mahmoud Al-Atiyatb, Agusto Luzuriaga-Neiraa, Osman Mahgoub

Gaafarc, Isam T. Kadimd, Waleed Al-Marzooqic, Hamza A. Babikere, Mohammed N. Al-Kindie,

Aliya S. Al-Ansarif, Ali H. Al-Lawatig, Albano Beja-Pereiraa,h

aResearch Center in Biodiversity and Genetic Resources (CIBIO), InBIO, University of Porto, R.

Padre Armando Quintas 7, 4485-661 Vairão, Porto, PortugalbDepartment of Animal Production, Faculty of Agriculture, Mutah University, Al-Karak 61710,

JordancDepartment of Animal and Veterinary Sciences, College of Agricultural and Marine Sciences,

Sultan Qaboos University, P.O. Box 34, Postal Code 123, Al-Khod, Sultanate of OmandDepartment of Biological Sciences and Chemistry, College of Arts and Sciences, University of

Nizwa, P.O. Box 33, Postal Code 616, Birkat Al-Mouz, Sultanate of OmaneDepartment of Biochemistry, College of Medicine and Health Sciences, Sultan Qaboos

University, P.O. Box 35, Postal Code 123, Al-Khod, Sultanate of OmanfDepartment of Biology, College of Sciences, Sultan Qaboos University, P.O. Box 36, Postal

Code 123, Al-Khod, Sultanate of OmangThe Oman Animal and Plant Genetic Resources Center, The Research Council, P.O. Box 1422,

Postal Code 130, Muscat, Sultanate of OmanhDepartment of Geosciences, Environment and Spatial Planning, Faculty of Sciences, University

of Porto, Rua Campo Alegre 687, 4169-007 Porto, Portugal

*Corresponding author

E-mail address: nsralaraimi@cibio.up.pt (N.A. Al-Araimi)

1

1

2

3

4

5

6

7

8

9

10

11

12

13

14

15

16

17

18

19

20

21

22

23

24

25

26

27

28

Supplementary tables

Table S1. General microsatellite markers used in this study.

Locus Origin Cr. a Multiplex Label Primer Sequences (5' - 3’) b Acc. no c ReferenceSRCRSP05 Caprine 21 MIX1 PET F: GGACTCTACCAACTGAGCTACAAG

R: TGAAATGAAGCTAAAGCAATGCL22197 Arevalo et al. (1994)

SRCRSP08* Caprine Unknown NED F: TGCGGTCTGGTTCTGATTTCACR: CCTGCATGAGAAAGTCGATGCTTAG

L22193 Bhebhe et al. (1994)

HSC* Ovine 20 VIC F: CTGCCAATGCAGAGACACAAGAR: GTCTGTCTCCTGTCTTGTCATC

M90759 Baumung et al. (2006)

MAF65 Ovine 15 VIC F: AAAGGCCAGAGTATGCAATTAGGAGR: CCACTCCTCCTGAGAATATAACATG

M67437 Buchanan et al. (1994)

INRA005 Bovine 12 FAM F: TTCAGGCATACCCTACACCACATGR: AAATATTAGCCAACTGAAAACTGGG

X63793 Vaiman et al. (1992)

INRA006* Bovine 3 PET F: AGGAATATCTGTATCAACCGCAGTCR: CTGAGCTGGGGTGGGAGCTATAAATA

X63793 Vaiman et al. (1992)

INRA063 Bovine 18 FAM F: GACCACAAAGGGATTTGCACAAGCR: AAACCACAGAAATGCTTGGAAG

X71507 Vaiman et al. (1994b)

ILSTS019 Bovine 21 FAM F: AGGGACCTCATGTAGAAGCR: ACTTTTGGACCCTGTAGTGC

L23492 Kemp et al. (1993b)

SRCRSP23 Caprine Unknown MIX2 FAM F: TGAACGGGTAAAGATGTGR: TGTTTTTAATGGCTGAGTAG

- Yeh et al. (1997)

ILSTS008 Ovine 9 VIC F: GAATCATGGATTTTCTGGGGR: TAGCAGTGAGTGAGGTTGGC

L23483 Kemp et al. (1993a)

CSRD247 Ovine 14 VIC F: GGACTTGCCAGAACTCTGCAATR: CACTGTGGTTTGTATTAGTCAGG

EU009450 Kemp et al. (1995)

McM527 Ovine 5 NED F: GTCCATTGCCTCAAATCAATTCR: AAACCACTTGACTACTCCCCAA

L34277 Crawford et al. (1995)

OarFCB20 Ovine 2 NED F: GGAAAACCCCCATATATACCTATACR: AAATGTGTTTAAGATTCCATACATGTG

L20004 Buchanan et al. (1994)

INRA023 Bovine 3 FAM F: GAGTAGAGCTACAAGATAAACTTCR: TAACTACAGGGTGTTAGATGAACTC

X67830 Vaiman et al. (1994a)

ILSTS087* Bovine 28 FAM F: AGCAGACATGATGACTCAGCR: CTGCCTCTTTTCTTGAGAGC

L37279 Kemp et al. (1995)

a Cr, chromosome location; b F and R, forward and reverse primer, respectively; c accession no; * loci were excluded from further analysis based on inconsistent banding patterns, HWE and linkage disequilibrium.

2

29

30

3132

Table S2. MICRO-CHECKER summary results of the presence of null alleles by population.

Locus/Population

Jabal Akhdar

Batinah Dhofar Ash

SharqiyahMusanda

mSomali

a Iran Pakistan India

SRCRSP05 no yes yes no no no no no noSRCRSP08 no no no no no no no yes noMAF65 no no no no no no no no noINRA005 no no no no no no no no noINRA006 no no no no no no yes no noINRA063 no no no no no no no no noILSTS19 no no yes no no no no no noSRCRSP23 no no no no no no no no noILSTS08 no no no no no no no no noCSRD247 no no no no no no no no noMcM527 no no no no no no no no noOarFCB20 no no no no yes no no no noINRA023 no no no no no no no no noILSTS87 no no no no no no no no no

MICRO-CHECKER summary results by population. The program highlighted the possible presence of null alleles in only six

instances (shown in bold yes).

3

33

34

35

36

Table S3. FST estimates with and without ENA correction.

Locus FST not using ENA FST using ENASRCRSP5 0.081 0.076SRCRSP8 0.211 0.202MAF65 0.089 0.087INRA005 0.053 0.053INRA006 0.054 0.055INRA063 0.117 0.115ILSTS19 0.140 0.137SRCRSP23 0.063 0.063ILSTS008 0.107 0.105CSRD247 0.107 0.106McM527 0.051 0.051OarFCB20 0.100 0.104INRA023 0.071 0.071ILSTS87 0.125 0.125All loci 0.096 0.095Bootstrap 95% CI 0.076-0.121 0.076-0.117

FreeNA output. The global FST (Weir and Cockerham, 1996) was estimated with and without ENA correction, as described in Chapuis

and Estoup (2007).

4

37

38

39

40

Table S4. Significant exact probability (P) of Hardy-Weinberg departures for each locus-population combination.

Locus/Population

Jabal Akhdar

Batinah

Dhofar

Ash Sharqiyah

Musandam

Somalia

Iran

Pakistan

India

HWEd ‡

SRCRSP5 no no no no no no no no no 0SRCRSP8 no no no no no no no no yes 1MAF65 no no no no no no no no no 0INRA005 no no no no no no no no no 0INRA006 no no no no no no yes no yes 2INRA063 no no no no no no no no no 0ILSTS19 no no no no no no no no no 0SRCRSP23 no no no no no no no no no 0ILSTS008 no no no no no no no no no 0CSRD247 no no no no no no no no no 0McM527 no no no no no no no no no 0OarFCB20 no no no no no no no no no 0INRA023 no no no no no no no no no 0ILSTS87 no no no no no no yes no no 1HWEd

‡ 0 0 0 0 0 0 2 0 2 0Bold yes indicates a locus-population combination that is still significant (P ≤ 0.001) following adjustments for multiple tests with

Holm–Bonferroni sequential correction (Holm, 1979); ‘no’ indicates not significant. HWEd ‡: No. of significant population and locus

departures from HWE at P ≤ 0.001 after Holm–Bonferroni sequential correction (Holm, 1979).

5

41

42

43

44

45

Table S5. Genotypic linkage disequilibrium tests. The number of significant (P < 0.001) locus combinations of 819 is listed for

each breed following adjustments for multiple tests with Holm–Bonferroni sequential correction (Holm, 1979).

Breed Number of Significant Locus Pairs

Locus

Jabal Akhdar - -Batinah - -

Dhofar 2 SRCRSP05-INRA006; MAF65-INRA006

Ash Sharqiyah

2 SRCRSP05-INRA006; SRCRSP5-INRA006

Musandam - -

Somalia 1 SRCRSP23-ILSTS087

Iran 8 SRCRSP23-CSRD247; SRCRSP23-McM527; SRCRSP23-INRA023; CSRD247-INRA023; SRCRSP23-ILSTS087; CSRD247-ILSTS087; McM527-ILSTS087; INRA023-ILSTS087

Pakistan - -

India 9 SRCRSP05-MAF65; SRCRSP08-MAF65; MAF65-INRA005; SRCRSP05-INRA006; SRCRSP08-INRA006; MAF65-INRA006; INRA006-INRA063; SRCRSP05-ILSTS019; MAF65-ILSTS019

Total 22

6

46

47

48

Table S6. Linkage disequilibrium (LD) detected between 14 pairs of loci at P < 0.001 after Holm–Bonferroni sequential

correction (Holm, 1979).

Pair-loci Holm-Bonferroni SignificanceSRCRSP05-MAF65 SignSRCRSP08-MAF65 SignSRCRSP05-INRA006 SignSRCRSP08-INRA006 SignMAF65-INRA006 SignINRA006-INRA063 SignMAF65-ILSTS019 SignSRCRSP23-ILSTS087 SignCSRD247-ILSTS087 SignMcM527-ILSTS087 SignMAF65-INRA005 SignSRCRSP05-ILSTS019 SignINRA006-ILSTS019 SignMAF65-INRA063 Sign

7

49

50

51

Table S7. Descriptive statistics of genetic diversity for 9 goat populations based on 11 genotyped microsatellite markers.

Locus Na a Ar b Par c Ho

d He e

SRCRSP05 12 6.852 0.164 0.694 0.749MAF65 17 8.226 0.315 0.767 0.777INRA005 4 3.151 0.001 0.623 0.551INRA063 5 4.128 0.000 0.702 0.662ILSTS019 7 4.615 0.007 0.526 0.565SRCRSP23 14 8.516 0.125 0.775 0.759ILSTS008 8 3.627 0.362 0.435 0.429CSRD247 10 6.194 0.082 0.699 0.729McM527 9 6.260 0.111 0.738 0.733OarFCB20 11 5.146 0.318 0.540 0.535INRA023 13 8.460 0.229 0.841 0.807Mean 10 5.925 0.156 0.667 0.663s.d. 3.924 1.96 0.134 0.123 0.124

a Number of observed alleles, b allelic richness, c private allelic richness, d observed heterozygosity, e expected heterozygosity

8

52

53

54

Table S8. Pairwise FST genetic distances for Omani breeds and commercial goat populations.

Pop. JKH BTN DHR SHR MSN SOM IRN PAK INDJKH 0BTN 0.062 0DHR 0.122 0.137 0SHR 0.116 0.113 0.095 0MSN 0.077 0.097 0.071 0.073 0SOM 0.117 0.113 0.058 0.084 0.062 0IRN 0.107 0.112 0.095 0.035 0.045 0.078 0PAK 0.090 0.068 0.064 0.092 0.081 0.089 0.078 0IND 0.103 0.129 0.098 0.072 0.056 0.101 0.028 0.080 0

P-value ˂ 0.05; goat population names correspond to Table 1.

9

55

56

57

Table S9. Analysis of molecular variance for Omani breeds and between Omani and commercial goat populations.

Source of Variation

d.f. Sum of Squares

Variance Components

Percentage of Variation

FST P-value

Omani goat breeds

Between breeds 4 100.605 0.383 9.71 0.097 0.000

Within breeds 277 986.874 3.525 90.29Total 281 1087.479 3.946

Nine goat populations

Between populations

8 176.926 0.351 8.78 0.0878

0.000

Within populations

465 1697.175 3.650 91.22

Total 473 1874.101 4.001

d.f.: degrees of freedom; P-value ˂ 0.05; P-values were obtained based on 10,000 permutations.

10

58

59

60

Supplementary figures

Fig. S1. Estimates of the null allele frequency using the EM method of Dempster et al. (1977) with FreeNA software. No loci

were excluded from the subsequent within-breed analyses due to the high frequency of potential null alleles (estimates of r greater

than or equal to 2).

11

61

62

63

64

65

Fig. S2. (A) Neighbor-joining tree of Nei’s standard distance matrix of 237 individual goats constructed based on 11

microsatellite loci. (B) Neighbor-Net based on the matrix of Nei’s standard genetic distance (Ds) between nine goat

populations. Each colored branch/semicarved line represents a cluster congruent with STRUCTURE analysis at K = 3 (see Fig. 1B).

Bootstrap support calculated from 1,000 replicates is shown when greater than 50%. The scale bars represent the pairwise distance

matrix between different individuals and populations. Abbreviations for the names of the goat population are listed in Table 1.

12

66

67

68

69

70

71

Fig. S3. Discriminant analyses of principal component (DAPC) scatterplots showing the genetic structure of nine goat

populations. Each dot represents a single animal, and populations are distinguished by different colors according to the legend.

Clusters are indicated based on STRUCTURE analysis at K = 3 (see Fig. 1B). For population codes, please refer to Table 1.

13

72

73

74

75

References

Arevalo, E., Holder, D.A., Derr, J.N., Bhebhe, E., Linn, R.A., Ruvuna, F., Davis, S.K., Taylor,

J.F., 1994. Caprine microsatellite dinucleotide repeat polymorphisms at the SR‐CRSP‐1,

SR‐CRSP‐2, SR‐CRSP‐3, SR‐CRSP‐41 and SR‐CRSP‐5 loci. Anim. Genet. 25, 202.

https://doi.org/10.1111/j.1365-2052.1994.tb00124.x.

Baumung, R., Cubric‐Curik, V., Schwend, K., Achmann, R., Sölkner, J., 2006. Genetic

characterisation and breed assignment in Austrian sheep breeds using microsatellite

marker information. J. Anim. Breed. Genet. 123, 265–271.

https://doi.org/10.1111/j.1439-0388.2006.00583.x.

Bhebhe, E., Kogi, J., Holder, D.A., Arevalo, E., Derr, J.N., Linn, R.A., Ruvuna, F., Davis, S.K.,

Taylor, J.F., 1994. Caprine microsatellite dinucleotide repeat polymorphisms at the SR‐CRSP‐6, SR‐CRSP‐7, SR‐CRSP‐8, SR‐CRSP‐9 and SR‐CRSP‐10 loci. Anim. Genet. 25,

203. https://doi.org/10.1111/j.1365-2052.1994.tb00125.x.

Buchanan, F.C., Galloway, S.M., Crawford, A.M., 1994. Ovine microsatellites at the OarFCB5,

OarFCB19, OarFCB20, OarFCB48, OarFCB129 and OarFCB226 loci. Anim. Genet. 25,

60. https://doi.org/10.1111/j.1365-2052.1994.tb00418.x.

Chapuis, M.-P., Estoup, A., 2007. Microsatellite null alleles and estimation of population

differentiation. Mol. Biol. Evol. 24, 621–631. https://doi.org/10.1093/molbev/msl191.

Crawford, A.M., Dodds, K.G., Ede, A.J., Pierson, C.A., Montgomery, G.W., Garmonsway, H.G.,

Beattie, A.E., Davies, K., Maddox, J.F., Kappes, S.W., 1995. An autosomal genetic

linkage map of the sheep genome. Genetics 140, 703–724.

14

76

77

78

79

80

81

82

83

84

85

86

87

88

89

90

91

92

93

94

95

96

Dempster, A.P., Laird, N.M., Rubin, D.B., 1977. Maximum likelihood from incomplete data via

the EM algorithm. J. Royal Statist. Soc. Ser. B Method. 39, 1–38.

https://doi.org/10.1111/j.2517-6161.1977.tb01600.x.

Holm, S., 1979. A simple sequentially rejective multiple test procedure. Scand. J. Statist. 6, 65–

70.

Kemp, S.J., Brezinsky, L., Teale, A.J., 1993a. ILSTS008: a polymorphic bovine microsatellite.

Anim. Genet. 24, 74. https://doi.org/10.1111/j.1365-2052.1993.tb00934.x.

Kemp, S.J., Brezinsky, L., Teale, A.J., 1993b. A panel of bovine, ovine and caprine polymorphic

microsatellites. Anim. Genet. 24, 363–365. https://doi.org/10.1111/j.1365-

2052.1993.tb00341.x.

Kemp, S.J., Hishida, O., Wambugu, J., Rink, A., Longeri, M.L., Ma, R.Z., Da, Y., Lewin, H.A.,

Barendse, W., Teale, A.J., 1995. A panel of polymorphic bovine, ovine and caprine

microsatellite markers. Anim. Genet. 26, 299–306. https://doi.org/10.1111/j.1365-

2052.1995.tb02663.x.

Vaiman, D., Imam-Ghali, M., Moazami-Goudarzi, K., Guérin, G., Grohs, C., Levéziel, H., Saïdi-

Mehtar, N., 1994a. Conservation of a syntenic group of microsatellite loci between cattle

and sheep. Mammal. Gen. 5, 310–314. https://doi.org/10.1007/BF00389547.

Vaiman, D., Mercier, D., Moazami-Goudarzi, K., Eggen, A., Ciampolini, R., Lépingle, A.,

Velmala, R., Kaukinen, J., Varvio, S.-L., Martin, P., 1994b. A set of 99 cattle

microsatellites: characterization, synteny mapping, and polymorphism. Mammal. Gen. 5,

288–297. https://doi.org/10.1007/BF00389543.

15

97

98

99

100

101

102

103

104

105

106

107

108

109

110

111

112

113

114

115

116

117

Vaiman, D., Osta, R., Mercier, D., Grohs, C., Leveziel, H., 1992. Characterization of five new

bovine dinucleotide repeats. Anim. Genet. 23, 537–541. https://doi.org/10.1111/j.1365-

2052.1992.tb00175.x.

Weir, B.S., Cockerham, C., 1996. Genetic Data Analysis II: Methods for Discrete Population

Genetic Data. Sinauer Assoc., Inc., Sunderland, MA.

Yeh, C.C., Kogi, J.K., Holder, M.T., Guerra, T.M., Davis, S.K., Taylor, J.F., 1997. Caprine

microsatellite dinucleotide repeat polymorphisms at the SR-CRSP21, SR-CRSP22, SR-

CRSP23, SR-CRSP24, SR-CRSP25, SR-CRSP26 and SR-CRSP27 loci. Anim. Genet.

28, 380–381.

16

118

119

120

121

122

123

124

125

126