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Forensic Science International: Genetics xxx (2011) xxxxxx

Contents lists available at ScienceDirect

Forensic Science International: Genetics

journal homepage: www.elsevier.com/locate/fsig

Forensic Population Genetics Short Communication

Genetic data for 17 Y-chromosomal STR loci in Macedonians in the

Republic of Macedonia
Zlatko Jakovski a,*, Ksenija Nikolova a, Renata Jankova-Ajanovska a, Damir Marjanovic b,c,
Naris Pojskic b, Biljana Janeska a
a
b
c

Institute of Forensic Medicine and Criminology, School of Medicine, University Ss. Cyril and Methodious, Skopje, Macedonia
Institute for Genetic Engineering and Biotechnology, Sarajevo, Bosnia and Herzegovina
Genos, Planinska 1, 10000 Zagreb, Croatia

A R T I C L E I N F O

A B S T R A C T

Article history:
Received 14 July 2010
Received in revised form 10 November 2010
Accepted 2 April 2011

The population data were obtained for the 16 Y chromosomal STR loci included in the AmpFistr1YlerTM
PCR Amplication Kit (DYS456, DYS389I, DYS390, DYS389II, DYS458, DYS19, DYS385 a/b, DYS393,
DYS391, DYS439, DYS635, DYS392, Y-GATA H4, DYS437, DYS438, DYS448) in a sample of 262 unrelated
men from the Republic of Macedonia.
2011 Elsevier Ireland Ltd. All rights reserved.

Keywords:
Y chromosomal short tandem repeats (STR)
Macedonian population data
AmpFistr1YlerTM PCR Amplication Kit

Populations: This population study was carried out to establish

a forensic database on the Macedonian population. The population
data were obtained for the 16 Y chromosomal STR loci included in
the AmpFistr1YlerTM PCR Amplication Kit (Applied Biosysetm)
(DYS456, DYS389I, DYS390, DYS389II, DYS458, DYS19, DYS385 a/b,
DYS393, DYS391, DYS439, DYS635, DYS392, Y-GATA H4, DYS437,
DYS438, DYS448). Similar population databases were found for
most countries in the area [18] (Fig. 1). The DNA samples
originate from 262 unrelated males. In view of the fact that the
Republic of Macedonia is a multi-national country with inhabitants of various ethnic origins and religions, it would be worth
noting that the foregoing males are Orthodox Christians of
Macedonian ethnicity from different geographical regions in the
Republic of Macedonia.
DNA extraction: Whole blood samples were collected on FTA
classic cards (Whatman) and stored at room temperature. DNA
from whole blood was extracted with QiaAmp DNA MiniKit
(QIAGEN Gmbh, Hilden Germany).
PCR: One multiplex PCR reaction was carried out for each DNA
sample using the AmpFistr1YlerTM PCR Amplication Kit
(Applied Biosysetm) according to the suppliers protocols. Amplication was carried out in a 9700 Thermal Cycler (Applied
Biosystems).
Typing: For electrophoresis, 1.5 ml of the PCR product was
combined with 12 ml of formamide and 0.5 ml of GeneScan 500 LIZ

* Corresponding author. Tel.: +389 2 3 177 0 44; fax: +389 2 1 178 831.
E-mail address: zlatedr@yahoo.com (Z. Jakovski).

size standard. Detection of PCR products and genotyping were

carried out on the ABI PRISM 310 Genetic Analyser (Applied
Biosystems) using the data collection software and GeneMapper ID
v3.2 analysis software (Applied Biosystems). The allele nomenclature is according to the Y-STR Haplotype Reference Database, YHRD
(Y-STR Haplotype Reference Database, http://www.yhrd.org).
Data analysis: An estimation of allele and haplotype frequency
distribution, haplotype and gene diversity was done using Neis
statistics [9]. An exact test of sample differentiation [10] based on
haplotype frequencies have been calculated in order to estimate a
possible difference among the Macedonian and the other observed
control populations. The considered signicance level after Bonferronis correction was p < 0.01. In order to determine the Macedonian
population Y_STR genetic relationships with the populations in the
neighboring regions (Bosnia and Herzegovina, Bulgaria, Croatia,
Kosovo, Serbia), share allele genetic distance [11] has been applied.
Also neighbor-joining dendrogram [12] based on result of genetic
distance analysis has been constructed. Consensus tree has been
constructed upon 1000 bootstraps. Principal components (PC)
analysis was performed on allele frequencies by using PAST software
[13]. Since only 7 same Y-STR loci were available from previous
studies for all included populations from neighboring regions,
DYS19, DYS389I, DYS389II, DYS390, DYS391, DYS392, DYS393, loci
have been considered during the genetic distance analysis.
The above mentioned calculations were made using Arlequin
version 3.11 [14], PowerMaker version 3.32 [15] and MEGA4 [16]
software.
Access to the data: Available upon request by email to:
zlatedr@yahoo.com.

1872-4973/$ see front matter 2011 Elsevier Ireland Ltd. All rights reserved.
doi:10.1016/j.fsigen.2011.04.005

Please cite this article in press as: Z. Jakovski, et al., Genetic data for 17 Y-chromosomal STR loci in Macedonians in the Republic of
Macedonia, Forensic Sci. Int. Genet. (2011), doi:10.1016/j.fsigen.2011.04.005

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Z. Jakovski et al. / Forensic Science International: Genetics xxx (2011) xxxxxx

e2

Fig. 1. Geographical location of Republic of Macedonia (red label) and other neighboring countries which population data were used (yellow label).

Results: Total of 243 different Y-STR haplotypes from 262

samples were detected: 227 of them were unique, 16 haplotypes
appeared twice and just one three times. An estimation of
allele frequencies distribution, the most frequent haplotypes

and haplotype diversity within the samples of the Macedonian

population are presented in Tables 1 and 2. Major allele
frequency and gene diversity across loci are presented in
Table 3.

Table 1
Allele frequency distribution of observed Y STR loci in the Macedonian population.
Marker

Allele

Count

Freq

Marker

Allele

Count

Freq

DYS19

13
14
15
16
17
11
12
13
14
9
27
28
29
30
31
32
33
21
22
23
24
25
26
10
11
12
9
10
11
12
13
14
16
10
12
13
14
15
10
11
12
13
14
15
15.3
16
16.3
17
18
19
20
21

73
54
64
56
15
1
43
180
37
1
1
31
47
103
61
16
3
2
15
52
156
36
1
162
86
2
12
2
222
11
25
1
1
1
60
185
13
3
3
62
9
43
118
70
2
88
1
50
61
11
4
2

0.2786
0.2061
0.2443
0.2137
0.0573
0.0038
0.1641
0.6870
0.1412
0.0038
0.0038
0.1183
0.1794
0.3931
0.2328
0.0611
0.0115
0.0076
0.0573
0.1985
0.5954
0.1374
0.0038
0.6183
0.3282
0.0076
0.0458
0.0076
0.8473
0.0420
0.0954
0.0038
0.0038
0.0038
0.2290
0.7061
0.0496
0.0115
0.0057
0.1183
0.0172
0.0821
0.2252
0.1336
0.0038
0.1679
0.0019
0.0954
0.1164
0.0210
0.0076
0.0038

27.9
20.6
24.4
21.4
5.7
0.4
16.4
68.7
14.1
0.4
0.4
11.8
17.9
39.3
23.3
6.1
1.1
0.8
5.7
19.8
59.5
13.7
0.4
61.8
32.8
0.8
4.6
0.8
84.7
4.2
9.5
0.4
0.4
0.4
22.9
70.6
5.0
1.1
0.6
11.8
1.7
8.2
22.5
13.4
0.4
16.8
0.2
9.5
11.6
2.1
0.8
0.4

DYS438

10
11
12
13
8
9
10
11
12
13
14
15
14
14.1
15
16
16
18
19
20
21
22
12
13
14
15
15.3
16
16.1
17
18
21
13
14
15
16
17
17.2
18
19
20
21
20
21
22
23
24
25
26
10
11
12
13
14
9

156
40
25
1
1
39
24
68
114
44
11
1
148
1
80
33
2
7
77
146
28
2
1
16
19
109
1
74
1
32
8
1
1
14
79
71
67
6
14
4
5
1
11
58
64
109
17
2
1
3
117
112
25
1
4

0.5954
0.1527
0.0954
0.0038
0.0038
0.1489
0.0916
0.2595
0.4351
0.1679
0.0420
0.0038
0.5649
0.0038
0.3053
0.1260
0.0076
0.0267
0.2939
0.5573
0.1069
0.0076
0.0038
0.0611
0.0725
0.4160
0.0038
0.2824
0.0038
0.1221
0.0305
0.0038
0.0038
0.0534
0.3015
0.2710
0.2557
0.0229
0.0534
0.0153
0.0191
0.0038
0.0420
0.2214
0.2443
0.4160
0.0649
0.0076
0.0038
0.0115
0.4466
0.4275
0.0954
0.0038
0.0153

59.5
15.3
9.5
0.4
0.4
14.9
9.2
26.0
43.5
16.8
4.2
0.4
56.5
0.4
30.5
12.6
0.8
2.7
29.4
55.7
10.7
0.8
0.4
6.1
7.3
41.6
0.4
28.2
0.4
12.2
3.1
0.4
0.4
5.3
30.2
27.1
25.6
2.3
5.3
1.5
1.9
0.4
4.2
22.1
24.4
41.6
6.5
0.8
0.4
1.1
44.7
42.7
9.5
0.4
1.5

DYS389I

DYS389II

DYS390

DYS391

DYS392

DYS393

DYS385 (a and b)

DYS439

DYS437

DYS448

DYS456

DYS458

DYS635

GATA-H4

Please cite this article in press as: Z. Jakovski, et al., Genetic data for 17 Y-chromosomal STR loci in Macedonians in the Republic of
Macedonia, Forensic Sci. Int. Genet. (2011), doi:10.1016/j.fsigen.2011.04.005

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Table 2
Haplotypes that have been detected more then once in the Macedonian population
sample and observed haplotype diversity.

e3

Table 3
Major allele frequency and gene diversity for observed Y STR loci in examined
sample of the Macedonian population.

Haplotypes

Count

Locus

Major allele freq.

Allele no.

Gene diversity

17-13-31-25-12-11-13-13-15-10-13-15-20-15-17-21-11
16-13-31-22-10-11-12-13-15-9-12-14-21-15-15-21-12
17-13-31-25-11-11-13-11-13-11-11-14-20-15-15-23-13
13-13-30-24-11-11-13-16-18-10-12-14-20-16-15-23-12
16-13-31-24-11-11-13-14-15-10-13-15-19-15-17-23-11
16-13-30-24-10-11-13-11-14-11-10-14-20-16-15-24-13
14-12-28-24-10-11-12-14-17-9-12-16-18-13-15-22-11
13-13-30-24-10-11-13-16-18-10-12-14-20-17-17-22-12
16-14-31-25-10-11-13-11-14-11-11-14-20-16-16-23-12
14-13-29-24-11-13-12-11-11-12-13-15-19-15-15-23-12
15-12-28-23-9-11-12-13-16-9-14-14-21-16-14-22-12
16-13-30-23-10-11-13-11-14-11-10-14-21-17-16-23-12
15-13-30-25-10-11-13-11-14-11-11-14-20-18-16-23-12
15-12-29-22-10-11-13-14-14-10-12-16-21-15-16-20-11
14-13-30-24-10-13-13-11-11-12-12-14-19-15-17-23-12
13-14-31-24-10-11-13-16-18-10-11-14-20-16-15-22-12
15-13-31-24-10-11-13-17-18-10-12-14-20-17-16-23-12
Haplotype diversity

2
2
2
2
2
2
2
3
2
2
2
2
2
2
2
2
2
1.0000  0.0003

DYS19
DYS389I
DYS389II
DYS390
DYS391
DYS392
DYS393
DYS385 a and b
DYS438
DYS439
DYS437
DYS448
DYS456
DYS458
DYS635
GATA-H4
Mean

0.2786
0.6870
0.3931
0.5954
0.6183
0.8473
0.7061
0.2252
0.5954
0.4351
0.5649
0.5573
0.4160
0.3015
0.4160
0.4466
0.5052

5
5
7
6
4
6
5
14
6
6
4
6
10
10
7
6
6.6875

0.7713
0.4811
0.7412
0.5839
0.5078
0.2711
0.4464
0.8590
0.5909
0.7049
0.5718
0.5908
0.7222
0.7634
0.7122
0.6084
0.6204

There are no statistical signicant differences in allele

frequencies between the Macedonian and the other observed
populations (p > 0.01).
The Macedonian population has the largest genetic distance
against the Croatian population (0.2643), while the lowest against
Bulgarian population (0.0815) (Table 4). The neighbor-joining
consensus tree constructed based on the results of the genetic
distance analysis shows that the population of the Macedonian
ethnic origin has possible highest similarity with the Bulgarian
populations, and not such dissimilarity with the cluster which
consists Bosnian and Herzegovinian-Serbian-Croatian populations,
while the Kosovo population create a separate group (Fig. 2).
The rst component PCA reveals that Macedonian population is
genetically closest to the Bulgarian and partially Serbian population especially compared with Kosovo population. The second
component PCA shows also closest relation of Macedonian and
Bulgarian populations. Croatian population is located at the other
end of the dimension since Bosnia and Herzeogina population has
separated position in compared with Macedonia and Bulgaria
populations. Kosovo population clusters separately among studied
populations. Total variance of the data set was 62%, from which the
rst and second PCs represented, 36% and 26% respectively (Fig. 3).
Conclusion: Previously published data for the Macedonian
population which were announced 5 years ago were based on the
observation of the 10 loci [1]. The main intention of this study was
to expand published Y-STR genetic data for the Macedonian
population with the information from the seven new loci, as well to
present some additional information about the Y-STR genetic
relationships of the Macedonian population with the human
populations from the neighboring countries.
According to the results of the comparative analysis, we can
conclude that population differences between the six observed
populations are expected. Macedonian population shows moderate Y-STR differentiation versus other Balkan populations. The

Table 4
Share allele distance among observed populations based on 7 observed Y STR loci.
Population

B&H

Bulgaria

Croatia

Kosovo

Macedonia

B&H
Bulgaria
Croatia
Kosovo
Macedonia
Serbia

0.1579
0.2036
0.2321
0.1376
0.1155

0.2687
0.1766
0.0815
0.1128

0.3291
0.2643
0.2160

0.1543
0.2064

0.1157

Serbia

percentage of unique haplotype (84.64%) is very high as well as

haplotype diversity. Gene diversity shows variation over loci in
range 0.27110.8590 with overall average value 0.6204.
The Balkan region includes human populations with high level
of Y STR diversity and with a quite similar genetic structure. It is
clear that the observed populations from central and eastern
Balkan do not have a pronounced differentiation of Y STR
population structure, except partially Kosovo population which
shows some of Y STR specicity. As the obtained results indicate,
genetic distance recorded between Macedonian population and
ethnic Albanians from Kosovo is relatively high. This nding is
curious considering that Kosovo and Republic Macedonia share a
border. In addition, almost 25% of Republic Macedonia citizens
constitute ethnic Albanians who share the origins with Albanian
population in Kosovo. Therefore, the results of this study suggest
that forensic experts in Macedonia need to apply caution when
estimating haplotype frequencies that are to be used in the
quantitative assessment of matches in forensic and genealogical
casework. Ethnicity of the examined individuals needs to be taken
into consideration and appropriate population database used in
these estimations.
In the course of the elaboration of this population study, we
adhered to the new guidelines published in this journal, wherein
we would like to point out that we accept the requirements

Kosovo
Bulgaria
68

96

Croatia
Bosnia and Herzegovina

92

Serbia
Macedonia

Fig. 2. Neighbor-joining consensus tree (1000 bootstraps) based on share allele genetic distance analyze estimated using the available data for 7 Y STR loci.

Please cite this article in press as: Z. Jakovski, et al., Genetic data for 17 Y-chromosomal STR loci in Macedonians in the Republic of
Macedonia, Forensic Sci. Int. Genet. (2011), doi:10.1016/j.fsigen.2011.04.005

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e4

1.6
1.2
Bulgaria

Component 2

0.8
Serbia
0.4
Macedonia
-2

-1.6
Croatia

-1.2

-0.8

-0.4
0.4
Bosnia_and_Herzegovina

0.8

1.2

1.6

-0.4
-0.8
-1.2
-1.6
Kosovo

-2
Component 1

Fig. 3. PC analysis performed using allele frequencies of available data for 7 Y STR loci.

References
[1] M. Spiroski, T. Arsov, C. Kruger, S. Willuweit, L. Roewer, Y-chromosomal STR
haplotypes in Macedonian population samples, Forensic Sci. Int. 148 (2005) 69
73.
[2] M. Pericic, I.K. Martinovic, L.L. Barac, B. Janicijevic, D. orpevic, L. Efremovska, P.
Rudan, Population genetics of 8 Y chromosome STR loci in Macedonians and
Macedonian Romani (Gypsy), Forensic Sci. Int. 154 (2005) 257261.
[3] M. Pericic, B.L. Lauc, I.K. Martinovic, B. Janicijevic, I. Behluli, P. Rudan, Y chromosome haplotypes in Albanian population from Kosovo, Forensic Sci. Int. 146
(2004) 6164.
[4] M. Stevanovic, V. Dobricic, D. Keckarevic, A. Perovic, D.S. Pavicevic, M.K. Markovic,
A. Jovanovic, S. Romac, Human Y-specic STR haplotypes in population of Serbia
and Montenegro, Forensic Sci. Int. 171 (2007) 216221.
[5] B.L. Lauc, M. Pericic, M.I. Klaric, A. Sijacki, D. Popovic, B. Janicijevic, P. Rudan, Y
chromosome STR polymorphisms in a Serbian population sample, Forensic Sci.
Int. 150 (2005) 97101.
[6] D. Marjanovic, N. Bakal, N. Pojskic, L. Kapur, K. Drobnic, D. Primorac, K. Bajrovic, R.
Hadziselimovic, Population data for the twelve Y-chromosome short tandem
repeat loci from the sample of multinational population in Bosnia and Herzegovina, J. Forensic Sci. 50 (2005) 223225.
[7] B. Zaharova, S. Andonova, A. Gilissen, J. Cassiman, R. Decorte, I. Kremensky, Ychromosomal STR haplotypes in three major population groups in Bulgaria,
Forensic Sci. Int. 124 (2001) 182186.

pertaining to the publication of this type of papers [17].

Furthermore, this paper is in line both with the original [18],
and the updated ISFG guidelines [19] of the DNA Commission of the
International Society of Forensic Genetics on the use of Y-STRs in
forensic analysis.
Quality control: GEDNAP (German DNA Proling Group) blind
trial system [20].

[8] L. Barac, M. Peric ic, I. Martinovic Klaric, B. Janicijevic, J. Parik, S. Rootsi, P. Rudan, Y
chromosome STRs in Croatians, Forensic Sci. Int. 138 (2003) 127133.
[9] M. Nei, Molecular Evolutionary Genetics, Columbia University Press, New York,
1987.
[10] M. Raymond, F. Rousset, An exact test for population differentiation, Evolution 49
(1995) 12801283.
[11] R. Chakraborty, L. Jin, Determination of relatedness between individuals using
DNA ngerprinting, Hum. Biol. 65 (1993) 875895.
[12] N. Saitou, M. Nei, The neighbor-joining method: a new method for reconstructing
phylogenetic trees, Mol. Biol. Evol. 4 (1987) 406425.
[13] R. Hammer, D.A.T. Harper, P.D. Ryan, PAST: Paleontological Statistics Software
Package for Education and Data Analysis, Palaeontologia Electronica 4 (2001) 19.
http://palaeo/electronica.org/2001_1/past/issue1_01.htm.
[14] L. Excofer, G. Laval, S. Schneider, Arlequin ver. 3.0: an integrated software
package for population genetics data analysis, Evol. Bioinform. Online 1 (2005)
4750.
[15] L. Kejun, V.M. Spencer, PowerMarker: an integrated analysis environment for
genetic marker analysis, Bioinformatics 21 (2005) 21282129.
[16] S. Kumar, J. Dudley, M. Nei, K. Tamura, MEGA: a biologist-centric software for
evolutionary analysis of DNA and protein sequences, Briengs Bioinform. 9
(2008) 299306.
[17] A. Carracedo, J.M. Butler, L. Gusmao, W. Parson, L. Roewer, P.M. Schneider,
Publication of population data for forensic purposes, Forensic Sci. Int. Genet. 4
(2010) 145147.
[18] P. Gill, C. Brenner, B. Brinkmann, B. Budowle, A. Carracedo, M.A. Jobling, P. de
Knijff, M. Kayser, M. Krawczak, W.R. Mayr, N. Morling, B. Olaisen, V. Pascali, M.
Prinz, L. Roewer, P.M. Schneider, A. Sajantila, C. Tyler-Smith, DNA Commission of
the International Society of Forensic Genetics: recommendations on forensic
analysis using Y-chromosome STRs, Forensic Sci. Int. 124 (2001) 510.
[19] L. Gusmao, J.M. Butler, A. Carracedo, P. Gill, M. Kayser, W.R. Mayr, N. Morling, M.
Prinz, L. Roewer, C. Tyler-Smith, P.M. Schneider, DNA Commission of the International Society of Forensic Genetics (ISFG): an update of the recommendations
on the use of Y-STRs in forensic analysis, Forensic Sci Int. 157 (2006) 187197.
[20] S. Rand, M. Schurenkamp, C. Hohoff, B. Brinkmann, The GEDNAP blind trial
concept part II, trends and developments, Int. J. Legal. Med. 118 (2004) 8389.

Please cite this article in press as: Z. Jakovski, et al., Genetic data for 17 Y-chromosomal STR loci in Macedonians in the Republic of
Macedonia, Forensic Sci. Int. Genet. (2011), doi:10.1016/j.fsigen.2011.04.005