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Genetic Polymorphisms of four X-STR Loci: DXS6797, DXS6800, HPRTB and GATA172D05 in a Peruvian population sample
Available online at www.sciencedirect.com
Forensic Science International: Genetics Supplement Series 1 (2008) 153–154 www.elsevier.com/locate/FSIGSS
Research article
Genetic Polymorphisms of four X-STR Loci: DXS6797, DXS6800, HPRTB and GATA172D05 in a Peruvian population sample R.E. Martinez a,b, M.L. Bravo c, D. Aguirre c, S.I. Polo a, D.V. Huerta a, L. Gusma˜o d, J.J. Builes c,e,* Laboratorio de Biologı´a Molecular y A´cidos Nucleicos, CIBN - UNMSM - Lima, Peru b ADN INSTITUTE, Lima, Peru c GENES Ltda. Medellı´n, Colombia d IPATIMUP, Instituto de Patologia e Imunologia Molecular da Universidade do Porto, R. Dr. Roberto Frias, s/n. 4200-465 Porto, Portugal e Instituto de Biologı´a, Universidad de Antioquia, Medellı´n, Colombia a
Received 22 August 2007; accepted 10 October 2007
Abstract The X linked short tandem repeats (STR) markers have proven to be very useful tools for paternity testing when the disputed child is female. The purpose of this study was to describe the genetic polymorphism of four X-chromosomal STR loci (DXS6797, DXS6800, HPRTB and GATA172D05) in a Peruvian population sample and evaluate their efficiency in forensic practice and paternity testing. # 2008 Elsevier Ireland Ltd. All rights reserved. Keywords: Forensic genetic; Peru; X-STR; DXS6797; DXS6800; HPRTB; GATA172D05; Population genetics
1. Introduction The X-STRs were recently recognized as important tools in forensic application, particularly in complex cases of kinship testing [1], especially in certain deficiency cases that can be solved by this technique rather than by using autosomal and Ychromosome (ChrY) testing [2]. The aim of this study was to describe and to analyze allele frequencies of four X-STR loci (DXS6797, DXS6800, HPRTB and GATA172D05) in a sample population of unrelated individual (mixed background) from Peru and add those markers to the panel of paternity testing and forensically used markers. 2. Material and methods Buccal swab samples were collected from 172 unrelated Peruvians (99 females and 73 males) obtaining consent to be involved in this study. Genomic DNA was extracted using the Salting out protocol [3]. * Corresponding author at: Genes Ltda, Carrera 48 No, 10-45 Cons. 611, Medellı´n, Colombia. Tel.: +57 4 268 48 75; fax: +57 4 318 52 70. E-mail addresses: [email protected], [email protected] (J.J. Builes). 1875-1768/$ – see front matter # 2008 Elsevier Ireland Ltd. All rights reserved. doi:10.1016/j.fsigss.2007.10.131
STRs were amplified in a multiplex system with primers of published sequences [4,8,9,10]. PCR was carried out in a 15 ml volume reaction containing 2–10 ng DNA, 1X Go Taq Buffer, 200 mM each dNTPs, 2.0 mM MgCl2, 0.5 U GoTaq, 2.4 mM DXS6797, 2.0 mM DXS6800, 2.0 mM HPRTB, 3.6 mM GATA172D05. Thermal cycling was conducted using PTC 100 (MJ Research). The PCR product were analyzed on 6% denaturing poliacrylamide gel electrophoresis containing 7 M urea and visualized by silver staining [5]. Alleles were identified based on the number of variable repeats and their attribution was made by comparison with an in-house constructed allelic ladder, DNA controls (including K562 and 9947A) and following the published nomenclature and ISFG guidelines for STR analysis [6]. Allelic frequencies were determined with software Arlequin Ver. 3.1 [7].
3. Results and discussion Allele frequencies of the examined X-linked STR in Peruvian samples are show in Table 1; allele frequency between male and female samples were not significantly different in all examined markers (date no show).
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R.E. Martinez et al. / Forensic Science International: Genetics Supplement Series 1 (2008) 153–154
Table 1 Allelic frequencies for the investigated loci in a sampled population of unrelated individual from Peru´
in Allele frequencies distribution of DXS6800 in Austria and Germany [13].
Allele
Conflict of interest
6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28
DXS 6797
0.0037 0.0037 0.0185 0.3284 0.2546 0.2731 0.0959 0.0185
DXS 6800
0.8007 0.0111 0.059 0.1181
HPRTB
0.0480 0.2066 0.4502 0.2177 0.0664 0.0111
GATA 172D05 0.1107 0.0037 0.0812 0.0369 0.3764 0.2915 0.0996
0.0111
0.0037
The Comparison of Allele frequencies distribution of DXS6797 in our sample is similar to Korean population [9], and Germans [11]; Allele frequencies distribution of GATA172D05 is similar to Spain population [12]; and different
None. References [1] J. Edelmann, R. Szibor, Electrophoresis 20 (1999) 2844–2846. [2] R. Szibor, M. Krawczak, S. Hering, J. Edelmann, E. Kuhlisch, D. Krause, Int. J. Leg. Med. 117 (2003) 67–74. [3] S. Miller, D. Dykes, H. Polesky, Nucleic Acids Res. 16 (1988) 1215. [4] J Murray, V Sheffield, J Weber, and G Duyk. Cooperative Human Linkage Center. 1995. http://www.ncbi.nlm.nih.gov/entrez/query.fcgicmd=retrieve.nucleotide/genbak. [5] C.J Sanguinetti, E Dias Neto, A.J Simpson.; 17(5) (1994) 914-21. [6] W. Ba¨r, B. Brinkmann, P. Lincoln, W.R. Rossi, B. Budowle, C. Bell, et al. Forensic Sci. Int. 87 (1997) 179–184. [7] P Lewis, D Zaykin. 2001. Genetic Data Analysis: computer program for the analysis of allelic data. Version 1.0 (d16c). Free Software on internet (http://lewis.eeb.uconn.edu/lewishome/software.html). [8] GDB: Genome Database (http://www.gdb.org). [9] S.H. Shin, J.S. Yu, S.W. Park, G.S. Min, K.W. Chung, Forensic Sci. Int. 147 (2005) 35–41. [10] J. Edelmann, D. Deichsel, S. Hering, I. Plate, R. Szibor, Forensic Sci. Int. 129 (2002) 99–103. [11] M. Poetsch, A. Repenning, E. Lignitz, E. Kuhlisch, R. Szibor, Int. J. Leg. Med. 120 (2006) 61–66. [12] M. Zarrabeitia, A. Alonso, J. Martı´n, A. Miguel, J. Martin-Escudero, et al. Int. J. Leg. Med. 120 (3) (2006) 147–150. [13] P. Wiegand, B. Berger, J. Edelmann, W. Parson, Int. J. Leg. Med. 117 (2003) 62–65.
Forensic Science International: Genetics Supplement Series 1 (2008) 153–154 www.elsevier.com/locate/FSIGSS
Research article
Genetic Polymorphisms of four X-STR Loci: DXS6797, DXS6800, HPRTB and GATA172D05 in a Peruvian population sample R.E. Martinez a,b, M.L. Bravo c, D. Aguirre c, S.I. Polo a, D.V. Huerta a, L. Gusma˜o d, J.J. Builes c,e,* Laboratorio de Biologı´a Molecular y A´cidos Nucleicos, CIBN - UNMSM - Lima, Peru b ADN INSTITUTE, Lima, Peru c GENES Ltda. Medellı´n, Colombia d IPATIMUP, Instituto de Patologia e Imunologia Molecular da Universidade do Porto, R. Dr. Roberto Frias, s/n. 4200-465 Porto, Portugal e Instituto de Biologı´a, Universidad de Antioquia, Medellı´n, Colombia a
Received 22 August 2007; accepted 10 October 2007
Abstract The X linked short tandem repeats (STR) markers have proven to be very useful tools for paternity testing when the disputed child is female. The purpose of this study was to describe the genetic polymorphism of four X-chromosomal STR loci (DXS6797, DXS6800, HPRTB and GATA172D05) in a Peruvian population sample and evaluate their efficiency in forensic practice and paternity testing. # 2008 Elsevier Ireland Ltd. All rights reserved. Keywords: Forensic genetic; Peru; X-STR; DXS6797; DXS6800; HPRTB; GATA172D05; Population genetics
1. Introduction The X-STRs were recently recognized as important tools in forensic application, particularly in complex cases of kinship testing [1], especially in certain deficiency cases that can be solved by this technique rather than by using autosomal and Ychromosome (ChrY) testing [2]. The aim of this study was to describe and to analyze allele frequencies of four X-STR loci (DXS6797, DXS6800, HPRTB and GATA172D05) in a sample population of unrelated individual (mixed background) from Peru and add those markers to the panel of paternity testing and forensically used markers. 2. Material and methods Buccal swab samples were collected from 172 unrelated Peruvians (99 females and 73 males) obtaining consent to be involved in this study. Genomic DNA was extracted using the Salting out protocol [3]. * Corresponding author at: Genes Ltda, Carrera 48 No, 10-45 Cons. 611, Medellı´n, Colombia. Tel.: +57 4 268 48 75; fax: +57 4 318 52 70. E-mail addresses: [email protected], [email protected] (J.J. Builes). 1875-1768/$ – see front matter # 2008 Elsevier Ireland Ltd. All rights reserved. doi:10.1016/j.fsigss.2007.10.131
STRs were amplified in a multiplex system with primers of published sequences [4,8,9,10]. PCR was carried out in a 15 ml volume reaction containing 2–10 ng DNA, 1X Go Taq Buffer, 200 mM each dNTPs, 2.0 mM MgCl2, 0.5 U GoTaq, 2.4 mM DXS6797, 2.0 mM DXS6800, 2.0 mM HPRTB, 3.6 mM GATA172D05. Thermal cycling was conducted using PTC 100 (MJ Research). The PCR product were analyzed on 6% denaturing poliacrylamide gel electrophoresis containing 7 M urea and visualized by silver staining [5]. Alleles were identified based on the number of variable repeats and their attribution was made by comparison with an in-house constructed allelic ladder, DNA controls (including K562 and 9947A) and following the published nomenclature and ISFG guidelines for STR analysis [6]. Allelic frequencies were determined with software Arlequin Ver. 3.1 [7].
3. Results and discussion Allele frequencies of the examined X-linked STR in Peruvian samples are show in Table 1; allele frequency between male and female samples were not significantly different in all examined markers (date no show).
154
R.E. Martinez et al. / Forensic Science International: Genetics Supplement Series 1 (2008) 153–154
Table 1 Allelic frequencies for the investigated loci in a sampled population of unrelated individual from Peru´
in Allele frequencies distribution of DXS6800 in Austria and Germany [13].
Allele
Conflict of interest
6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28
DXS 6797
0.0037 0.0037 0.0185 0.3284 0.2546 0.2731 0.0959 0.0185
DXS 6800
0.8007 0.0111 0.059 0.1181
HPRTB
0.0480 0.2066 0.4502 0.2177 0.0664 0.0111
GATA 172D05 0.1107 0.0037 0.0812 0.0369 0.3764 0.2915 0.0996
0.0111
0.0037
The Comparison of Allele frequencies distribution of DXS6797 in our sample is similar to Korean population [9], and Germans [11]; Allele frequencies distribution of GATA172D05 is similar to Spain population [12]; and different
None. References [1] J. Edelmann, R. Szibor, Electrophoresis 20 (1999) 2844–2846. [2] R. Szibor, M. Krawczak, S. Hering, J. Edelmann, E. Kuhlisch, D. Krause, Int. J. Leg. Med. 117 (2003) 67–74. [3] S. Miller, D. Dykes, H. Polesky, Nucleic Acids Res. 16 (1988) 1215. [4] J Murray, V Sheffield, J Weber, and G Duyk. Cooperative Human Linkage Center. 1995. http://www.ncbi.nlm.nih.gov/entrez/query.fcgicmd=retrieve.nucleotide/genbak. [5] C.J Sanguinetti, E Dias Neto, A.J Simpson.; 17(5) (1994) 914-21. [6] W. Ba¨r, B. Brinkmann, P. Lincoln, W.R. Rossi, B. Budowle, C. Bell, et al. Forensic Sci. Int. 87 (1997) 179–184. [7] P Lewis, D Zaykin. 2001. Genetic Data Analysis: computer program for the analysis of allelic data. Version 1.0 (d16c). Free Software on internet (http://lewis.eeb.uconn.edu/lewishome/software.html). [8] GDB: Genome Database (http://www.gdb.org). [9] S.H. Shin, J.S. Yu, S.W. Park, G.S. Min, K.W. Chung, Forensic Sci. Int. 147 (2005) 35–41. [10] J. Edelmann, D. Deichsel, S. Hering, I. Plate, R. Szibor, Forensic Sci. Int. 129 (2002) 99–103. [11] M. Poetsch, A. Repenning, E. Lignitz, E. Kuhlisch, R. Szibor, Int. J. Leg. Med. 120 (2006) 61–66. [12] M. Zarrabeitia, A. Alonso, J. Martı´n, A. Miguel, J. Martin-Escudero, et al. Int. J. Leg. Med. 120 (3) (2006) 147–150. [13] P. Wiegand, B. Berger, J. Edelmann, W. Parson, Int. J. Leg. Med. 117 (2003) 62–65.