- Email: [email protected]
Population genetic analysis of 12 X-STRs in Swedish population
Forensic Science International: Genetics 6 (2012) e80–e81
Contents lists available at ScienceDirect
Forensic Science International: Genetics journal homepage: www.elsevier.com/locate/fsig
Forensic population genetics—Letter to the Editor Population genetic analysis of 12 X-STRs in Swedish population
Dear Editor, DNA markers on the X-chromosome have been shown to be a powerful tool for solving complex relationship cases [1,2]. The main application of X-chromosomal markers is in deficient paternity cases, especially the investigations of multiple females with the hypothesis that they share the same father [3]. Two different concepts of genetic properties, in addition to population frequencies, become relevant to study when using multiple markers from the same chromosome. These features include physical dependency between loci (genetic linkage) and the allelic dependency between alleles at different loci (linkage disequilibrium (LD), or gametic disequilibrium) [4]. Genetic linkage can be studied by observing the degree of recombination fraction between markers within families, and LD can be studied by comparing observed haplotype frequencies with expected haplotype frequencies inferred from allele frequencies based on data from unrelated individuals. Here we report on data from 652 unrelated males from a Swedish population that were typed for the 12 X-chromosomal Short Tandem Repeat (STR) markers included in the Investigator Argus X-12 kit (Qiagen). The markers are located in four different linkage groups (linkage group 1: DXS10148, DXS10135, DXS8378; linkage group 2: DXS7132, DXS10079, DXS10074; linkage group 3: DXS10103, HPRTB, DXS10101; linkage group 4: DXS10146, DXS10134, DXS7423). The samples come from blood donors and have previously been shown to be good representatives of the Swedish population [5]. In addition, 107 individuals from 17 Swedish families were typed for the estimation of recombination fractions. In these families, four to seven children were accompanied by a genetically confirmed mother and father (paternity/ maternity index > 10,000, based on autosomal STRs). All samples were analysed with the Investigator Argus X-12 Kit according to the manufacturer’s instructions (Qiagen). PCRamplicons were analysed on an ABI Prism 3100 capillary electrophoresis sequencer (Applied Biosystems) using the filter set recommended by the manufacturer and with the sequenced allelic ladder included in the kit as reference. Alleles were binned using GeneMapper ID ver. 3.2 with the binset provided by the manufacturer (Qiagen). The typing of the 652 Swedish males resulted in 652 different haplotypes when all 12 X-STRs were included (Supplementary Table 1 and 2). Associations between alleles from different loci were tested using the Genetic Data Analysis software program by Lewis and Zaykin [6]. In this test a comparison was performed between pairs of loci and P-values were obtained using exact test with 10,000 permutations. The test revealed significant P-values (P < 0.0008 after Bonferroni correction) for pair of markers within the linkage groups but not between 1872-4973/$ – see front matter ß 2011 Elsevier Ireland Ltd. All rights reserved. doi:10.1016/j.fsigen.2011.07.008
different linkage groups (Supplementary Table 3). These results strengthen the fact that allelic dependencies exist within linkage groups, thus suggesting the use of observed haplotypes frequencies rather than expected haplotype frequencies inferred from allele frequencies. When examining the population data as a group of markers (i.e. linkage group), linkage group 1 was shown to be the most polymorfic group with 367 different observed haplotypes (Supplementary Tables 2 and 4) and consequently the group with the highest informativeness when studying different forensic efficiency parameters (Supplementary Table 2). The least informative group of markers was linkage group 3 with 195 different haplotypes among the 652 individuals tested. In this group the most common haplotype was seen 24 times, which corresponds to a population frequency of 3.7%. Only minor, non-significant, differences (measured by FSTvalues) were observed when the Swedish haplotype frequencies were compared with corresponding frequencies for the same set of markers in a German population [7] and in a Hungarian population sample [8] (Supplementary Table 5). Due to the close physical location of the markers and the fact that all markers are located on the same chromosome it is of utmost importance to examine the linkage situation for this set of markers. We estimated the recombination fraction between nearby located loci with the method described earlier by Tillmar et al. [9]. In total, using data from 74 theoretical meioses, we showed that there is a low probability for recombinations to occur within a linkage group. For the families tested, recombinations within linkage groups were only observed in linkage group 2, between markers DXS7132 and DXS10079 (Supplementary Table 6). For the recombination fractions between linkage groups, linkage groups 3 and 4 gave an estimate of 0.28, with a 95% credibility interval not including 0.5. However, larger studies need to be performed in order to get more precise estimates for the closely located markers within each linkage group. As earlier suggested [9–12] both linkage and LD should be accounted for when using this, and similar sets of markers. It is, however, difficult to measure the impact of the observed linkage situation in forensic casework. One way to present the effect of the found LD, is to compare likelihood ratios (LRs) computed with expected haplotype frequencies inferred from allele frequencies with LRs computed with frequencies from haplotype count for a relevant pedigree. For such purpose, we performed a simulation study for a deficiency case where two females were tested whether or not they have the same father, and when DNA-data is missing for the alleged father [9]. DNA-data for 10,000 such cases where simulated, and for each case LR was calculated with both observed haplotype frequencies and haplotype frequencies inferred from allele frequencies. The two LRs were compared to measure the case specific effect of ignoring LD. The simulation showed that the LR would generally be overestimated ignoring the degree of LD found
Letter to the Editor / Forensic Science International: Genetics 6 (2012) e80–e81
in the Swedish population sample using this set of the 12 X-STRs (Supplementary Fig. 1). We conclude that using Investigator Argus X-12 in forensic deficiency cases can be useful and powerful, and that linkage and LD should be accounted for when using this set of markers for the Swedish population. This paper follows the guidelines for publication of population data as requested by the journal [13]. Acknowledgement We would like to acknowledge Helena Nilsson for technical assistance. Appendix A. Supplementary data Supplementary data associated with this article can be found, in the online version, at doi:10.1016/j.fsigen.2011.07.008. References [1] R. Szibor, S. Hering, E. Kuhlisch, I. Plate, S. Demberger, M. Krawczak, J. Edelmann, Haplotyping of STR cluster DXS6801-DXS6809-DXS6789 on Xq21 provides a powerful tool for kinship testing, Int. J. Legal Med. 119 (2005) 363–369. [2] R. Szibor, M. Krawczak, S. Hering, J. Edelmann, E. Kuhlisch, D. Krause, Use of Xlinked markers for forensic purposes, Int. J. Legal Med. 117 (2003) 67–74. [3] R. Szibor, X-chromosomal markers: past, present and future, Forensic Sci. Int. Genet. 1 (2007) 93–99. [4] J. Ott, Analysis of Human Genetic Linkage, 3rd ed., The Johns Hopkins University Press, Baltimore, 1999. [5] A.O. Karlsson, T. Wallerstro¨m, A. Go¨therstro¨m, G. Holmlund, Y-chromosome diversity in Sweden—a long-time perspective, Eur. J. Hum. Genet. 14 (2006) 963–970.
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[6] P.O. Lewis, D. Zaykin, Genetic Data Analysis: Computer Program for the Analysis of Allelic Data, 2001, p. 1.1. [7] J. Edelmann, S. Lutz-Bonengel, J. Naue, S. Hering, X-chromosomal haplotype frequencies of four linkage groups using the Investigator Argus X-12 Kit, Forensic Sci. Int. Genet. (2011), doi:10.1016/j.fsigen.2011.01.001. [8] G. Horva´th, A. Zala´n, Z. Kis, H. Pamjav, A genetic study of 12 X-STR loci in the Hungarian population, Forensic Sci. Int. Genet. (2011), doi:10.1016/j.fsigen.2011.03.007. [9] A.O. Tillmar, P. Mostad, T. Egeland, B. Lindblom, G. Holmlund, K. Montelius, Analysis of linkage and linkage disequilibrium for eight X-STR markers, Forensic Sci. Int. Genet. 3 (2008) 37–41. [10] A.O. Tillmar, T. Egeland, B. Lindblom, G. Holmlund, P. Mostad, Using X-chromosomal markers in relationship testing: calculation of likelihood ratios taking both linkage and linkage disequilibrium into account, Forensic Sci. Int. Genet. (2010), doi:10.1016/j.fsigen.2010.11.00. [11] E. Medina-Acosta, F.B. Machado, Eyes wide open: the (mis)use of combined power of discrimination for X-linked short tandem repeats, Mol. Biol. Rep. 38 (2011) 4003–4006. [12] S. Inturri, S. Menegon, A. Amoroso, C. Torre, C. Robino, Linkage and linkage disequilibrium analysis of X-STRs in Italian families, Forensic Sci. Int. Genet. 5 (2011) 152–154. [13] A. Carracedo, J.M. Butler, L. Gusmao, W. Parson, L. Roewer, P. Schneider, Publication of population data for forensic purposes, Forensic Sci. Int. Genet. 4 (2010) 145–147.
Andreas O. Tillmar* National Board of Forensic Medicine, Department of Forensic Genetics and Forensic Toxicology, Artillerigatan 12, SE-58758 Linko¨ping, Sweden *Tel.: +46 13 25 21 43; fax: +46 13 13 60 05 E-mail address: [email protected] (A.O. Tillmar).
6 May 2011
Contents lists available at ScienceDirect
Forensic Science International: Genetics journal homepage: www.elsevier.com/locate/fsig
Forensic population genetics—Letter to the Editor Population genetic analysis of 12 X-STRs in Swedish population
Dear Editor, DNA markers on the X-chromosome have been shown to be a powerful tool for solving complex relationship cases [1,2]. The main application of X-chromosomal markers is in deficient paternity cases, especially the investigations of multiple females with the hypothesis that they share the same father [3]. Two different concepts of genetic properties, in addition to population frequencies, become relevant to study when using multiple markers from the same chromosome. These features include physical dependency between loci (genetic linkage) and the allelic dependency between alleles at different loci (linkage disequilibrium (LD), or gametic disequilibrium) [4]. Genetic linkage can be studied by observing the degree of recombination fraction between markers within families, and LD can be studied by comparing observed haplotype frequencies with expected haplotype frequencies inferred from allele frequencies based on data from unrelated individuals. Here we report on data from 652 unrelated males from a Swedish population that were typed for the 12 X-chromosomal Short Tandem Repeat (STR) markers included in the Investigator Argus X-12 kit (Qiagen). The markers are located in four different linkage groups (linkage group 1: DXS10148, DXS10135, DXS8378; linkage group 2: DXS7132, DXS10079, DXS10074; linkage group 3: DXS10103, HPRTB, DXS10101; linkage group 4: DXS10146, DXS10134, DXS7423). The samples come from blood donors and have previously been shown to be good representatives of the Swedish population [5]. In addition, 107 individuals from 17 Swedish families were typed for the estimation of recombination fractions. In these families, four to seven children were accompanied by a genetically confirmed mother and father (paternity/ maternity index > 10,000, based on autosomal STRs). All samples were analysed with the Investigator Argus X-12 Kit according to the manufacturer’s instructions (Qiagen). PCRamplicons were analysed on an ABI Prism 3100 capillary electrophoresis sequencer (Applied Biosystems) using the filter set recommended by the manufacturer and with the sequenced allelic ladder included in the kit as reference. Alleles were binned using GeneMapper ID ver. 3.2 with the binset provided by the manufacturer (Qiagen). The typing of the 652 Swedish males resulted in 652 different haplotypes when all 12 X-STRs were included (Supplementary Table 1 and 2). Associations between alleles from different loci were tested using the Genetic Data Analysis software program by Lewis and Zaykin [6]. In this test a comparison was performed between pairs of loci and P-values were obtained using exact test with 10,000 permutations. The test revealed significant P-values (P < 0.0008 after Bonferroni correction) for pair of markers within the linkage groups but not between 1872-4973/$ – see front matter ß 2011 Elsevier Ireland Ltd. All rights reserved. doi:10.1016/j.fsigen.2011.07.008
different linkage groups (Supplementary Table 3). These results strengthen the fact that allelic dependencies exist within linkage groups, thus suggesting the use of observed haplotypes frequencies rather than expected haplotype frequencies inferred from allele frequencies. When examining the population data as a group of markers (i.e. linkage group), linkage group 1 was shown to be the most polymorfic group with 367 different observed haplotypes (Supplementary Tables 2 and 4) and consequently the group with the highest informativeness when studying different forensic efficiency parameters (Supplementary Table 2). The least informative group of markers was linkage group 3 with 195 different haplotypes among the 652 individuals tested. In this group the most common haplotype was seen 24 times, which corresponds to a population frequency of 3.7%. Only minor, non-significant, differences (measured by FSTvalues) were observed when the Swedish haplotype frequencies were compared with corresponding frequencies for the same set of markers in a German population [7] and in a Hungarian population sample [8] (Supplementary Table 5). Due to the close physical location of the markers and the fact that all markers are located on the same chromosome it is of utmost importance to examine the linkage situation for this set of markers. We estimated the recombination fraction between nearby located loci with the method described earlier by Tillmar et al. [9]. In total, using data from 74 theoretical meioses, we showed that there is a low probability for recombinations to occur within a linkage group. For the families tested, recombinations within linkage groups were only observed in linkage group 2, between markers DXS7132 and DXS10079 (Supplementary Table 6). For the recombination fractions between linkage groups, linkage groups 3 and 4 gave an estimate of 0.28, with a 95% credibility interval not including 0.5. However, larger studies need to be performed in order to get more precise estimates for the closely located markers within each linkage group. As earlier suggested [9–12] both linkage and LD should be accounted for when using this, and similar sets of markers. It is, however, difficult to measure the impact of the observed linkage situation in forensic casework. One way to present the effect of the found LD, is to compare likelihood ratios (LRs) computed with expected haplotype frequencies inferred from allele frequencies with LRs computed with frequencies from haplotype count for a relevant pedigree. For such purpose, we performed a simulation study for a deficiency case where two females were tested whether or not they have the same father, and when DNA-data is missing for the alleged father [9]. DNA-data for 10,000 such cases where simulated, and for each case LR was calculated with both observed haplotype frequencies and haplotype frequencies inferred from allele frequencies. The two LRs were compared to measure the case specific effect of ignoring LD. The simulation showed that the LR would generally be overestimated ignoring the degree of LD found
Letter to the Editor / Forensic Science International: Genetics 6 (2012) e80–e81
in the Swedish population sample using this set of the 12 X-STRs (Supplementary Fig. 1). We conclude that using Investigator Argus X-12 in forensic deficiency cases can be useful and powerful, and that linkage and LD should be accounted for when using this set of markers for the Swedish population. This paper follows the guidelines for publication of population data as requested by the journal [13]. Acknowledgement We would like to acknowledge Helena Nilsson for technical assistance. Appendix A. Supplementary data Supplementary data associated with this article can be found, in the online version, at doi:10.1016/j.fsigen.2011.07.008. References [1] R. Szibor, S. Hering, E. Kuhlisch, I. Plate, S. Demberger, M. Krawczak, J. Edelmann, Haplotyping of STR cluster DXS6801-DXS6809-DXS6789 on Xq21 provides a powerful tool for kinship testing, Int. J. Legal Med. 119 (2005) 363–369. [2] R. Szibor, M. Krawczak, S. Hering, J. Edelmann, E. Kuhlisch, D. Krause, Use of Xlinked markers for forensic purposes, Int. J. Legal Med. 117 (2003) 67–74. [3] R. Szibor, X-chromosomal markers: past, present and future, Forensic Sci. Int. Genet. 1 (2007) 93–99. [4] J. Ott, Analysis of Human Genetic Linkage, 3rd ed., The Johns Hopkins University Press, Baltimore, 1999. [5] A.O. Karlsson, T. Wallerstro¨m, A. Go¨therstro¨m, G. Holmlund, Y-chromosome diversity in Sweden—a long-time perspective, Eur. J. Hum. Genet. 14 (2006) 963–970.
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[6] P.O. Lewis, D. Zaykin, Genetic Data Analysis: Computer Program for the Analysis of Allelic Data, 2001, p. 1.1. [7] J. Edelmann, S. Lutz-Bonengel, J. Naue, S. Hering, X-chromosomal haplotype frequencies of four linkage groups using the Investigator Argus X-12 Kit, Forensic Sci. Int. Genet. (2011), doi:10.1016/j.fsigen.2011.01.001. [8] G. Horva´th, A. Zala´n, Z. Kis, H. Pamjav, A genetic study of 12 X-STR loci in the Hungarian population, Forensic Sci. Int. Genet. (2011), doi:10.1016/j.fsigen.2011.03.007. [9] A.O. Tillmar, P. Mostad, T. Egeland, B. Lindblom, G. Holmlund, K. Montelius, Analysis of linkage and linkage disequilibrium for eight X-STR markers, Forensic Sci. Int. Genet. 3 (2008) 37–41. [10] A.O. Tillmar, T. Egeland, B. Lindblom, G. Holmlund, P. Mostad, Using X-chromosomal markers in relationship testing: calculation of likelihood ratios taking both linkage and linkage disequilibrium into account, Forensic Sci. Int. Genet. (2010), doi:10.1016/j.fsigen.2010.11.00. [11] E. Medina-Acosta, F.B. Machado, Eyes wide open: the (mis)use of combined power of discrimination for X-linked short tandem repeats, Mol. Biol. Rep. 38 (2011) 4003–4006. [12] S. Inturri, S. Menegon, A. Amoroso, C. Torre, C. Robino, Linkage and linkage disequilibrium analysis of X-STRs in Italian families, Forensic Sci. Int. Genet. 5 (2011) 152–154. [13] A. Carracedo, J.M. Butler, L. Gusmao, W. Parson, L. Roewer, P. Schneider, Publication of population data for forensic purposes, Forensic Sci. Int. Genet. 4 (2010) 145–147.
Andreas O. Tillmar* National Board of Forensic Medicine, Department of Forensic Genetics and Forensic Toxicology, Artillerigatan 12, SE-58758 Linko¨ping, Sweden *Tel.: +46 13 25 21 43; fax: +46 13 13 60 05 E-mail address: [email protected] (A.O. Tillmar).
6 May 2011