- Email: [email protected]
Pentaplex typing of new European Standard Set (ESS) STR loci in Indian population
Forensic Science International: Genetics 6 (2012) e86–e89
Contents lists available at ScienceDirect
Forensic Science International: Genetics journal homepage: www.elsevier.com/locate/fsig
Letter to the Editor Pentaplex typing of new European Standard Set (ESS) STR loci in Indian population
Dear Editor, Autosomal short tandem repeats (STRs) are effective tools for human identification. The amplicon size of the conventional STR markers used for the forensic casework ranges from 75 to 450 bp. Due to fragmented DNA template or PCR inhibitors present in the samples, often loss of signal or partial profiles are observed for the conventional STR markers. Previous studies have demonstrated utility of size-reduced amplicons (miniSTRs) for the analysis of highly degraded DNA by choosing primers close to the repeat regions [1–4]. European DNA profiling group (EDNAP) recommended five new European Standard Set (ESS) markers D10S1248, D2S441, D22S1045, D1S1656 and D12S391 for the forensic casework and database [5,6]. Evaluating polymorphisms of these miniSTR markers in the Indian population would enable their efficient use for the forensic casework. Hence, a pentaplex for the simultaneous analysis of these five miniSTR loci was optimized and their polymorphisms was studied in 11 endogamous populations of India belonging to four linguistic (Indo-European, Dravidian, Tibeto-Burman and Austro-Asiatic), six geographic (north, east, central, west, south and north-east) and two socio-cultural (caste and tribe) groups. 750 blood samples from unrelated male individuals belonging to eleven endogamous populations: Balmiki (62), Sakaldiwipi Brahmin (65), Kanyakubja Brahmin (78), Konkanastha Brahmin (71), Mahadev Koli (65), Iyengar (67), Kurumans (67), Gond (75), Tripuri (65), Riang (67) and Munda (68) were collected with informed consent following the protocols approved by the Institutional Ethnic Committee. Details of populations and there geographical, linguistic and social affinities have been described elsewhere [7]. Genomic DNA from the blood samples was isolated by the phenol-chloroform organic extraction method and quantified using the Quantifiler1 Human DNA Quantification kit according to the manufacturer’s protocol employing 7500 RealTime PCR System (Applied Biosystems, Foster city, CA). Pentaplex was built around ‘miniplex 01’ of Coble and Butler [3] replacing marker D14S1434 with D2S441 of ‘miniplex 02’. Amplification primers for the loci D1S1656 and D12S391 generating short amplicons were selected from the previous published papers [8–10]. Pentaplex primers were checked by the AutoDimer software for the primer–dimer and hairpin structures [11]. Spatial layout of markers showed four overlapping amplicons and the 50 end of the forward primers were labeled with 6-FAMTM (Blue), VICTM (Green), NEDTM (Yellow) and PETTM (Red) (Applied Biosystems, Foster city, CA) dyes for multiplexing. Dye labels of the primers and matrix files were identical with commonly used multiplexes in the forensic laboratories. Details of the loci and dye labels of pentaplex loci are listed in Table 1 (supplementary data). 1872-4973/$ – see front matter ß 2011 Elsevier Ireland Ltd. All rights reserved. doi:10.1016/j.fsigen.2011.07.003
Approximately 1 ng of DNA was amplified for pentaplex loci in a single reaction in GeneAmp1 PCR System 9700 (Applied Biosystems, Foster City, CA) in 9600 emulation mode (ramp speed of 1 8C/ s), following the amplification conditions which have been described elsewhere [3]. The multiplex was optimized by adjusting primer concentrations to balance peak heights within and between the fluorescent dye colours. The amplified products were electrophoresed in ABI PRISM1 3100 Genetic Analyzer with POP-4TM polymer using GeneScanTM 500 LIZTM as size standard. The data was analyzed using GeneMapper1 ID Software v3.2 (Applied Biosystems, Foster City, CA). The amplified products from the different samples were pooled together for generating allelic ladder. Four alleles for the each locus and all micro-variants were sequenced to confirm the repeat number, structure, and also to refine the bins and panels. Heterozygous alleles were separated by gel extraction method and sequenced in ABI PRISM1 3100 Genetic Analyzer using Big Dye1 Terminator v3.1 Cycle Sequencing kit, POP-6TM polymer (Applied Biosystems, Foster City, CA) and published primers [3,8–10]. DNA from the cell lines 9947A, 9948 and K562 (Promega Corp., USA) were used as standard reference materials. Sensitivity of pentaplex was studied by varying the template DNA quantity from 0.05 to 2 ng in 10 ml reaction. Optimum amplification for the pentaplex loci was achieved at an annealing temperature of 55 8C following the PCR conditions recommended by Coble and Butler [3]. The multiplex worked well with DNA quantities ranging from 0.2 to 1.0 ng with 0.5 ng DNA being the optimal template quantity. Due to the amplicon size in the range of 78–160 bp, pentaplex genotyping of degraded samples (four decomposed long bones and two teeth), recovered from wooded area (post mortem interval about 18–24 months), was processed. The replicate samples belonging to different individuals were pulverized at low temperature to obtain fine powder. DNA isolation was performed by organic extraction procedure followed by purification/concentration by Microcon1 100. Five decomposed tissues (muscles), recovered after few weeks, and were also processed. Pentaplex profiles observed from all replicate samples were in consensus and better than conventional STRs. Low copy number (LCN) typing has been a tool for low template DNA samples, but requires precautions and scientific skill to avoid mistakes in reading data [12–14]. Genetic profiles from templates less than 100 pg of DNA could not be developed by conventional STRs, especially touch DNA and degraded bones. Few such templates were processed with pentaplex genotyping procedure and partial profiles were observed from samples having more than 20 pg DNA. LCN typing for pentaplex was attempted by addition of AmpliTaq Gold1 DNA polymerase to the amplified product. With a pre-hold at 95 8C for 11 min, the products were amplified for 6 additional cycles. Full amplification resulting in peak height could be achieved by increasing the number of PCR cycles to 34 (Figs. 1 and 2). Negative controls and reagent blanks were tested to check
Letter to the Editor / Forensic Science International: Genetics 6 (2012) e86–e89
e87
Fig. 1. Pentaplex electrophoretogram without LCN.
the contamination and all the amplification reactions were conducted twice to assess consistency. Allele frequencies for the autosomal pentaplex loci, observed heterozygosity (H), gene diversity (GD), polymorphism information content (PIC), p values for the likelihood ratio and exact tests for the possible deviation from Hardy–Weinberg equilibrium were computed by PowerMarker program v3.25 [15]. Population pairwise genetic distances (Fst) were calculated using Arlequin programme v3.5 [16]. Forensic statistical parameters like probability of match (PM), power of discrimination (PD), power of exclusion (PE) and typical paternity index (TPI) were computed by PowerStats Excel macro v1.2 (Promega Corp., USA). Allele frequencies and values for different statistical parameters for the 11 populations are given in the Tables 2–6 (supplementary data). Tri-allelic genotype (18, 19, 23) pattern of ‘Type II’ (three peaks with even height) and rare allele 14 were observed at locus D12S391 in one sample each of Riang and Iyengar populations, respectively. Allele 7 and 19 were obtained for the FAMTM labeled loci D1S1656 and D22S1045 in a sample of the Balmiki and Gond populations, respectively which were confirmed by performing single-plex PCR reaction. High heterozygosity values ranging from 0.9487 to 0.7612 were obtained for all the studied miniSTR loci. The average observed heterozygosities of the selected loci were higher than the expected heterozygosities, except for the locus D2S441 in the studied populations. The studied populations were also genotyped for the 15 autosomal STR loci using the commercially available AmpFlSTR1 Identifiler kit. Comparison of heterozygosity values obtained for the miniSTR and Identifiler loci has been mentioned in Table 7 (supplementary data). The miniSTR locus D1S1656 was found to be the most polymorphic and
heterozygous among all loci with highest heterozygosity value of 0.9487, followed by D12S391 and FGA loci with comparable heterozygosity value (0.9104). The other three markers showed average polymorphisms in the studied populations. No deviations from Hardy–Weinberg equilibrium were observed after the application of Bonferroni correction (p < 0.01) in the studied populations. The combined PD, PE, PM and most common profile frequency for the studied populations are given in Table 8 (supplementary data). The combined PD values ranged from 0.999997741 to 0.999981885 in the Kanyakubja Brahmin and Riang populations, respectively. Combined PM values ranged from 1.81 105 in the Riang population to 9.65 106 in the Munda population. The combined powers of exclusion varied among the populations from 0.997310618 to 0.959422368 in the Kurumans and Riang populations, respectively. The most common profile frequencies for the populations ranged from 1.25 104 in the Mahadev Koli population to 8.52 105 in the Gond population. Results for the population pairwise genetic distances (Fst) among the studied populations are presented in Table 9 (supplementary data). The two TB populations (Tripuri and Riang) showed significant genetic distances from the remaining populations. This may be explained by the geographic confinement of TB populations to the north-eastern part of India, which is a landlocked hilly area preventing the genetic exchange between TB speakers and mainland Indian populations. The four Brahmin populations (Sakaldwipi Brahmin, Kanyakubja Brahmin, Konkanastha Brahmin and Iyengar) showed less genetic distances from each other, though they belong to different geographical and linguistic groups. This may be due to the forceful introduction of IE speakers into the pre-existing DR society and subsequent
e88
Letter to the Editor / Forensic Science International: Genetics 6 (2012) e86–e89
Fig. 2. Pentaplex electrophoretogram with LCN.
admixture between them [17]. The high genetic diversity among castes and tribes of India may be due to their different population histories and diverse geographical locations [18]. The present study attempts to create a pentaplex for the next generation markers and to survey the genetic diversity among few Indian populations using these reduced-size amplicons. The five miniSTR markers showed high heterozygosities, comparable to that obtained for the Identifiler loci in the studied populations. In addition to the usefulness of these markers in degraded DNA analysis like mass disasters, they might also complement the conventional markers in the deficiency cases. The data generated in the present study will help in constructing database of miniSTRs for Indian populations for forensic purposes and further studies in population genetics. CFSL regularly participates in the external proficiency testing programmes. This paper follows the guidelines and ISFG recommendations regarding the quality for publication of population data [19,20]. The study was funded by Directorate of Forensic Science Services, Ministry of Home Affairs, Govt. of India, New Delhi. The authors are thankful to all volunteers who participated in the study. Thanks are also to Mr Subhankar Nath, Dr Varsha Rajesh Rathod, Dr Mukesh Kumar Thakar and Dr Ganga Nath Jha of different institutes for their co-operation.
Appendix A. Supplementary data Supplementary data associated with this article can be found, in the online version, at doi:10.1016/j.fsigen.2011.07.003.
References [1] J.M. Butler, Y. Shen, B.R. McCord, The development of reduced size STR amplicons as tools for analysis of degraded DNA, J. Forensic Sci. 48 (2003) 1054–1064. [2] D.T. Chung, J. Drabek, K.L. Opel, J.M. Butler, B.R. McCord, A study on the effects of degradation and template concentration on the amplification efficiency of the STR miniplex primer sets, J. Forensic Sci. 49 (2004) 733–740. [3] M.D. Coble, J.M. Butler, Characterization of new miniSTR loci to aid analysis of degraded DNA, J. Forensic Sci. 50 (2005) 43–53. [4] C.R. Hill, M.C. Kline, M.D. Coble, J.M. Butler, Characterization of 26 miniSTR loci for improved analysis of degraded DNA samples, J. Forensic Sci. 53 (2008) 73–80. [5] P. Gill, L. Fereday, N. Morling, P.M. Schneider, New multiplexes for Europe— amendments and clarification of strategic development, Forensic Sci. Int. 163 (2006) 155–157. [6] P. Gill, L. Fereday, N. Morling, P.M. Schneider, The evolution of DNA databasesrecommendations for new European STR loci, Forensic Sci. Int. 156 (2006) 242–244. [7] T. Ghosh, D. Kalpana, S. Mukerjee, M. Mukherjee, A.K. Sharma, S. Nath, V.R. Rathod, M.K. Thakar, G.N. Jha, Genetic diversity of autosomal STRs in eleven populations of India, Forensic Sci. Int. Genet. 5 (2010) 259–261. , doi:10.1016/j.fsigen.2010.01.005. [8] M.V. Lareu, S. Barral, A. Salas, C. Pestoni, A. Carracedo, Sequence variation of a hypervariable short tandem repeat at the D1S1656 locus, Int. J. Legal Med. 111 (1998) 244–247. [9] M.V. Lareu, C. Pestoni, M. Schu¨renkamp, S. Rand, B. Brinkmann, A. Carracedo, A highly variable STR at the D12S391 locus, Int. J. Legal Med. 109 (1996) 134–138. [10] H. Asamura, S. Fujimori, M. Ota, H. Fukushima, MiniSTR multiplex systems based on non-CODIS loci for analysis of degraded DNA samples, Forensic Sci. Int. 173 (2007) 7–15. [11] P.M. Vallone, J.M. Butler, AutoDimer: a screening tool for primer–dimer and hairpin structures, Biotechniques 37 (2004) 226–231. [12] P. Gill, J. Whitaker, C. Flaxman, N. Brown, J. Buckleton, An investigation of the rigor of interpretation rules for STRs derived from less than 100 pg of DNA, Forensic Sci. Int. 112 (2000) 17–40.
Letter to the Editor / Forensic Science International: Genetics 6 (2012) e86–e89 [13] J.P. Whitaker, E.A. Cotton, P. Gill, A comparison of the characteristics of profiles produced with the AmpFlSTR SGM Plus multiplex system for both standard and low copy number (LCN) STR DNA analysis, Forensic Sci. Int. 123 (2001) 215–223. [14] P. Gill, Application of low copy number DNA profiling, Croat. Med. J. 42 (2001) 229–232. [15] J. Liu, S.V. Muse, PowerMarker: integrated analysis environment for genetic marker data, Bioinformatics 21 (2005) 2128–2129. [16] L. Excoffier, G. Lavel, S. Schneider, Arlequin ver 3.1: an integrated software package for population genetics data analysis, Evol. Bioinform. Online 1 (2005) 47–50. [17] M. Bamshad, T. Kivisild, W.S. Watkins, M.E. Dixon, C.E. Ricker, B.B. Rao, et al., Genetic evidence on the origins of Indian caste populations, Genome Res. 11 (2001) 994–1004. [18] P.P. Majumder, The human genetic history of south Asia, Curr. Biol. 23 (2010) 184–187. [19] B. Olaisen, W. Ba¨r, B. Brinkmann, B. Budowle, A. Carracedo, P. Gill, P. Lincoln, W.R. Mayr, S. Rand, DNA recommendations 1997 of the International Society of Forensic Genetics, Vox Sang. 74 (1998) 61–63. [20] 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) 145–147.
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D. Kalpana Tania Ghosh Sanjukta Mukerjee Meeta Mukherjee Anil Kumar Sharma* Central Forensic Science Laboratory, Directorate of Forensic Science Services, Ministry of Home Affairs, Govt. of India, 30 Gorachand Road, Park Circus, Kolkata 700014, West Bengal, India *Corresponding
author. Tel.: +91 33 22841638; fax: +91 33 22849442 E-mail address: [email protected] (A.K. Sharma). 23 November2010
Contents lists available at ScienceDirect
Forensic Science International: Genetics journal homepage: www.elsevier.com/locate/fsig
Letter to the Editor Pentaplex typing of new European Standard Set (ESS) STR loci in Indian population
Dear Editor, Autosomal short tandem repeats (STRs) are effective tools for human identification. The amplicon size of the conventional STR markers used for the forensic casework ranges from 75 to 450 bp. Due to fragmented DNA template or PCR inhibitors present in the samples, often loss of signal or partial profiles are observed for the conventional STR markers. Previous studies have demonstrated utility of size-reduced amplicons (miniSTRs) for the analysis of highly degraded DNA by choosing primers close to the repeat regions [1–4]. European DNA profiling group (EDNAP) recommended five new European Standard Set (ESS) markers D10S1248, D2S441, D22S1045, D1S1656 and D12S391 for the forensic casework and database [5,6]. Evaluating polymorphisms of these miniSTR markers in the Indian population would enable their efficient use for the forensic casework. Hence, a pentaplex for the simultaneous analysis of these five miniSTR loci was optimized and their polymorphisms was studied in 11 endogamous populations of India belonging to four linguistic (Indo-European, Dravidian, Tibeto-Burman and Austro-Asiatic), six geographic (north, east, central, west, south and north-east) and two socio-cultural (caste and tribe) groups. 750 blood samples from unrelated male individuals belonging to eleven endogamous populations: Balmiki (62), Sakaldiwipi Brahmin (65), Kanyakubja Brahmin (78), Konkanastha Brahmin (71), Mahadev Koli (65), Iyengar (67), Kurumans (67), Gond (75), Tripuri (65), Riang (67) and Munda (68) were collected with informed consent following the protocols approved by the Institutional Ethnic Committee. Details of populations and there geographical, linguistic and social affinities have been described elsewhere [7]. Genomic DNA from the blood samples was isolated by the phenol-chloroform organic extraction method and quantified using the Quantifiler1 Human DNA Quantification kit according to the manufacturer’s protocol employing 7500 RealTime PCR System (Applied Biosystems, Foster city, CA). Pentaplex was built around ‘miniplex 01’ of Coble and Butler [3] replacing marker D14S1434 with D2S441 of ‘miniplex 02’. Amplification primers for the loci D1S1656 and D12S391 generating short amplicons were selected from the previous published papers [8–10]. Pentaplex primers were checked by the AutoDimer software for the primer–dimer and hairpin structures [11]. Spatial layout of markers showed four overlapping amplicons and the 50 end of the forward primers were labeled with 6-FAMTM (Blue), VICTM (Green), NEDTM (Yellow) and PETTM (Red) (Applied Biosystems, Foster city, CA) dyes for multiplexing. Dye labels of the primers and matrix files were identical with commonly used multiplexes in the forensic laboratories. Details of the loci and dye labels of pentaplex loci are listed in Table 1 (supplementary data). 1872-4973/$ – see front matter ß 2011 Elsevier Ireland Ltd. All rights reserved. doi:10.1016/j.fsigen.2011.07.003
Approximately 1 ng of DNA was amplified for pentaplex loci in a single reaction in GeneAmp1 PCR System 9700 (Applied Biosystems, Foster City, CA) in 9600 emulation mode (ramp speed of 1 8C/ s), following the amplification conditions which have been described elsewhere [3]. The multiplex was optimized by adjusting primer concentrations to balance peak heights within and between the fluorescent dye colours. The amplified products were electrophoresed in ABI PRISM1 3100 Genetic Analyzer with POP-4TM polymer using GeneScanTM 500 LIZTM as size standard. The data was analyzed using GeneMapper1 ID Software v3.2 (Applied Biosystems, Foster City, CA). The amplified products from the different samples were pooled together for generating allelic ladder. Four alleles for the each locus and all micro-variants were sequenced to confirm the repeat number, structure, and also to refine the bins and panels. Heterozygous alleles were separated by gel extraction method and sequenced in ABI PRISM1 3100 Genetic Analyzer using Big Dye1 Terminator v3.1 Cycle Sequencing kit, POP-6TM polymer (Applied Biosystems, Foster City, CA) and published primers [3,8–10]. DNA from the cell lines 9947A, 9948 and K562 (Promega Corp., USA) were used as standard reference materials. Sensitivity of pentaplex was studied by varying the template DNA quantity from 0.05 to 2 ng in 10 ml reaction. Optimum amplification for the pentaplex loci was achieved at an annealing temperature of 55 8C following the PCR conditions recommended by Coble and Butler [3]. The multiplex worked well with DNA quantities ranging from 0.2 to 1.0 ng with 0.5 ng DNA being the optimal template quantity. Due to the amplicon size in the range of 78–160 bp, pentaplex genotyping of degraded samples (four decomposed long bones and two teeth), recovered from wooded area (post mortem interval about 18–24 months), was processed. The replicate samples belonging to different individuals were pulverized at low temperature to obtain fine powder. DNA isolation was performed by organic extraction procedure followed by purification/concentration by Microcon1 100. Five decomposed tissues (muscles), recovered after few weeks, and were also processed. Pentaplex profiles observed from all replicate samples were in consensus and better than conventional STRs. Low copy number (LCN) typing has been a tool for low template DNA samples, but requires precautions and scientific skill to avoid mistakes in reading data [12–14]. Genetic profiles from templates less than 100 pg of DNA could not be developed by conventional STRs, especially touch DNA and degraded bones. Few such templates were processed with pentaplex genotyping procedure and partial profiles were observed from samples having more than 20 pg DNA. LCN typing for pentaplex was attempted by addition of AmpliTaq Gold1 DNA polymerase to the amplified product. With a pre-hold at 95 8C for 11 min, the products were amplified for 6 additional cycles. Full amplification resulting in peak height could be achieved by increasing the number of PCR cycles to 34 (Figs. 1 and 2). Negative controls and reagent blanks were tested to check
Letter to the Editor / Forensic Science International: Genetics 6 (2012) e86–e89
e87
Fig. 1. Pentaplex electrophoretogram without LCN.
the contamination and all the amplification reactions were conducted twice to assess consistency. Allele frequencies for the autosomal pentaplex loci, observed heterozygosity (H), gene diversity (GD), polymorphism information content (PIC), p values for the likelihood ratio and exact tests for the possible deviation from Hardy–Weinberg equilibrium were computed by PowerMarker program v3.25 [15]. Population pairwise genetic distances (Fst) were calculated using Arlequin programme v3.5 [16]. Forensic statistical parameters like probability of match (PM), power of discrimination (PD), power of exclusion (PE) and typical paternity index (TPI) were computed by PowerStats Excel macro v1.2 (Promega Corp., USA). Allele frequencies and values for different statistical parameters for the 11 populations are given in the Tables 2–6 (supplementary data). Tri-allelic genotype (18, 19, 23) pattern of ‘Type II’ (three peaks with even height) and rare allele 14 were observed at locus D12S391 in one sample each of Riang and Iyengar populations, respectively. Allele 7 and 19 were obtained for the FAMTM labeled loci D1S1656 and D22S1045 in a sample of the Balmiki and Gond populations, respectively which were confirmed by performing single-plex PCR reaction. High heterozygosity values ranging from 0.9487 to 0.7612 were obtained for all the studied miniSTR loci. The average observed heterozygosities of the selected loci were higher than the expected heterozygosities, except for the locus D2S441 in the studied populations. The studied populations were also genotyped for the 15 autosomal STR loci using the commercially available AmpFlSTR1 Identifiler kit. Comparison of heterozygosity values obtained for the miniSTR and Identifiler loci has been mentioned in Table 7 (supplementary data). The miniSTR locus D1S1656 was found to be the most polymorphic and
heterozygous among all loci with highest heterozygosity value of 0.9487, followed by D12S391 and FGA loci with comparable heterozygosity value (0.9104). The other three markers showed average polymorphisms in the studied populations. No deviations from Hardy–Weinberg equilibrium were observed after the application of Bonferroni correction (p < 0.01) in the studied populations. The combined PD, PE, PM and most common profile frequency for the studied populations are given in Table 8 (supplementary data). The combined PD values ranged from 0.999997741 to 0.999981885 in the Kanyakubja Brahmin and Riang populations, respectively. Combined PM values ranged from 1.81 105 in the Riang population to 9.65 106 in the Munda population. The combined powers of exclusion varied among the populations from 0.997310618 to 0.959422368 in the Kurumans and Riang populations, respectively. The most common profile frequencies for the populations ranged from 1.25 104 in the Mahadev Koli population to 8.52 105 in the Gond population. Results for the population pairwise genetic distances (Fst) among the studied populations are presented in Table 9 (supplementary data). The two TB populations (Tripuri and Riang) showed significant genetic distances from the remaining populations. This may be explained by the geographic confinement of TB populations to the north-eastern part of India, which is a landlocked hilly area preventing the genetic exchange between TB speakers and mainland Indian populations. The four Brahmin populations (Sakaldwipi Brahmin, Kanyakubja Brahmin, Konkanastha Brahmin and Iyengar) showed less genetic distances from each other, though they belong to different geographical and linguistic groups. This may be due to the forceful introduction of IE speakers into the pre-existing DR society and subsequent
e88
Letter to the Editor / Forensic Science International: Genetics 6 (2012) e86–e89
Fig. 2. Pentaplex electrophoretogram with LCN.
admixture between them [17]. The high genetic diversity among castes and tribes of India may be due to their different population histories and diverse geographical locations [18]. The present study attempts to create a pentaplex for the next generation markers and to survey the genetic diversity among few Indian populations using these reduced-size amplicons. The five miniSTR markers showed high heterozygosities, comparable to that obtained for the Identifiler loci in the studied populations. In addition to the usefulness of these markers in degraded DNA analysis like mass disasters, they might also complement the conventional markers in the deficiency cases. The data generated in the present study will help in constructing database of miniSTRs for Indian populations for forensic purposes and further studies in population genetics. CFSL regularly participates in the external proficiency testing programmes. This paper follows the guidelines and ISFG recommendations regarding the quality for publication of population data [19,20]. The study was funded by Directorate of Forensic Science Services, Ministry of Home Affairs, Govt. of India, New Delhi. The authors are thankful to all volunteers who participated in the study. Thanks are also to Mr Subhankar Nath, Dr Varsha Rajesh Rathod, Dr Mukesh Kumar Thakar and Dr Ganga Nath Jha of different institutes for their co-operation.
Appendix A. Supplementary data Supplementary data associated with this article can be found, in the online version, at doi:10.1016/j.fsigen.2011.07.003.
References [1] J.M. Butler, Y. Shen, B.R. McCord, The development of reduced size STR amplicons as tools for analysis of degraded DNA, J. Forensic Sci. 48 (2003) 1054–1064. [2] D.T. Chung, J. Drabek, K.L. Opel, J.M. Butler, B.R. McCord, A study on the effects of degradation and template concentration on the amplification efficiency of the STR miniplex primer sets, J. Forensic Sci. 49 (2004) 733–740. [3] M.D. Coble, J.M. Butler, Characterization of new miniSTR loci to aid analysis of degraded DNA, J. Forensic Sci. 50 (2005) 43–53. [4] C.R. Hill, M.C. Kline, M.D. Coble, J.M. Butler, Characterization of 26 miniSTR loci for improved analysis of degraded DNA samples, J. Forensic Sci. 53 (2008) 73–80. [5] P. Gill, L. Fereday, N. Morling, P.M. Schneider, New multiplexes for Europe— amendments and clarification of strategic development, Forensic Sci. Int. 163 (2006) 155–157. [6] P. Gill, L. Fereday, N. Morling, P.M. Schneider, The evolution of DNA databasesrecommendations for new European STR loci, Forensic Sci. Int. 156 (2006) 242–244. [7] T. Ghosh, D. Kalpana, S. Mukerjee, M. Mukherjee, A.K. Sharma, S. Nath, V.R. Rathod, M.K. Thakar, G.N. Jha, Genetic diversity of autosomal STRs in eleven populations of India, Forensic Sci. Int. Genet. 5 (2010) 259–261. , doi:10.1016/j.fsigen.2010.01.005. [8] M.V. Lareu, S. Barral, A. Salas, C. Pestoni, A. Carracedo, Sequence variation of a hypervariable short tandem repeat at the D1S1656 locus, Int. J. Legal Med. 111 (1998) 244–247. [9] M.V. Lareu, C. Pestoni, M. Schu¨renkamp, S. Rand, B. Brinkmann, A. Carracedo, A highly variable STR at the D12S391 locus, Int. J. Legal Med. 109 (1996) 134–138. [10] H. Asamura, S. Fujimori, M. Ota, H. Fukushima, MiniSTR multiplex systems based on non-CODIS loci for analysis of degraded DNA samples, Forensic Sci. Int. 173 (2007) 7–15. [11] P.M. Vallone, J.M. Butler, AutoDimer: a screening tool for primer–dimer and hairpin structures, Biotechniques 37 (2004) 226–231. [12] P. Gill, J. Whitaker, C. Flaxman, N. Brown, J. Buckleton, An investigation of the rigor of interpretation rules for STRs derived from less than 100 pg of DNA, Forensic Sci. Int. 112 (2000) 17–40.
Letter to the Editor / Forensic Science International: Genetics 6 (2012) e86–e89 [13] J.P. Whitaker, E.A. Cotton, P. Gill, A comparison of the characteristics of profiles produced with the AmpFlSTR SGM Plus multiplex system for both standard and low copy number (LCN) STR DNA analysis, Forensic Sci. Int. 123 (2001) 215–223. [14] P. Gill, Application of low copy number DNA profiling, Croat. Med. J. 42 (2001) 229–232. [15] J. Liu, S.V. Muse, PowerMarker: integrated analysis environment for genetic marker data, Bioinformatics 21 (2005) 2128–2129. [16] L. Excoffier, G. Lavel, S. Schneider, Arlequin ver 3.1: an integrated software package for population genetics data analysis, Evol. Bioinform. Online 1 (2005) 47–50. [17] M. Bamshad, T. Kivisild, W.S. Watkins, M.E. Dixon, C.E. Ricker, B.B. Rao, et al., Genetic evidence on the origins of Indian caste populations, Genome Res. 11 (2001) 994–1004. [18] P.P. Majumder, The human genetic history of south Asia, Curr. Biol. 23 (2010) 184–187. [19] B. Olaisen, W. Ba¨r, B. Brinkmann, B. Budowle, A. Carracedo, P. Gill, P. Lincoln, W.R. Mayr, S. Rand, DNA recommendations 1997 of the International Society of Forensic Genetics, Vox Sang. 74 (1998) 61–63. [20] 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) 145–147.
e89
D. Kalpana Tania Ghosh Sanjukta Mukerjee Meeta Mukherjee Anil Kumar Sharma* Central Forensic Science Laboratory, Directorate of Forensic Science Services, Ministry of Home Affairs, Govt. of India, 30 Gorachand Road, Park Circus, Kolkata 700014, West Bengal, India *Corresponding
author. Tel.: +91 33 22841638; fax: +91 33 22849442 E-mail address: [email protected] (A.K. Sharma). 23 November2010