Development and validation of a next generation STR ESS-pentaplex

We constructed a simple STR pentaplex of new loci recommended as next generation markers for the European Standard Set (ESS) comprising normal-amplicon STRs: D12S391 and D1S1656, plus mini-amplicon STRs: D2S441, D10S1248 and D22S1045. Validation of the pentaplex included evaluation of its ability to amplify DNA from a variety of degraded forensic casework samples. Although the ESS-pentaplex was designed in the first instance to generate allele frequency data to supplement existing databases of established STRs, the multiplex proved to be a valuable tool for the analysis of challenging DNA when certain markers of Identifiler or MiniFiler occasionally failed.     

武汉汉族人群STR基因座D3S1358、D13S317、D12S391的多态性研究

目的　调查中国武汉地区汉族人群STR基因座—D3S1 358、D1 3S31 7、D1 2S391基因频率分布和群体遗传数据。方法　从 2 0 8个汉族无关个体收集血液标本 ,应用PCR技术及聚丙烯酰胺凝胶垂直板电泳对D3S1 358、D1 3S31 7和D1 2S391基因座分型。结果　D3S1 358检出 7个等位基因和 4 1个基因型。三基因座基因型分布符合Hardy-Weinberg平衡。观察 2 31次减数分裂均未发现突变基因。另外 ,调查结果计算显示D3S1 358、D1 3S31 7和D1 2S391基因座的杂合度 (H)分别为 0 70 98、 0 80 56和 0 84 0 0 ;三个人识别能力 (DP)分别为 0 851 6、 0 9332和 0 952 3;非父排除率 ( pE)分别为 0 4 463、 0 60 1 6和 0 681 8。结论　D3S1 358、D1 3S31 7和D1 2S391基因座在群体遗传学研究和法医学亲子鉴定及个人识别中具有较高实用价值     

Validation of nine non-CODIS STR loci for forensic use in a population from Central Poland

The D7S1517, D3S1744, D12S391, D2S1360, D6S474, D8S1132, D5S2500, D10S2325 and D4S236613 are STR loci potentially useful for forensic purposes whose analysis has recently become facilitated by availability of a commercial kit. The purpose of the study was to evaluate the usefulness of these loci for forensic identification in a population of Central Poland. The distribution of alleles of the nine STRs was determined in sample of 353 unrelated individuals born in Central Poland and indices of forensic informativeness were calculated. The studied loci were highly informative and did not show departures from Hardy-Weinberg equilibrium. For the loci located on the same chromosomes (D2S1360, D3S1744 D4S2366, D5S2500, D7S1517, D8S1132, D12S391) as other loci commonly used for identification purposes (TPOX, D2S1338, D3S1358, FGA, D5S818, D7S820, D8S1179 and D12S391) appropriate pairwise analysis of linkage disequilibrium was performed. In all cases no statistically significant deviation from independence was found. We conclude that the studied STRs are informative and, when necessary, can be used to extend the results obtained with other STRs commonly analyzed for identification purposes, in particular the CODIS set.     

Population data on the D1S1656 and D12S391 STR loci in Andalusia (south Spain) and the maghreb (north Africa).

Allele and genotype frequencies for two tetrameric short tandem repeat (STR) loci were determined in two population samples, from Andalusia (S Spain; n = 127) and the Maghreb (N Africa; n = 40). After denaturing polyacrylamide gel electrophoresis, 14 alleles were identified for D12S391 and 13 alleles for D1S1656. No deviations from the Hardy-Weinberg equilibrium were detected. Some statistical parameters of forensic interest (H, PD, EC) were also calculated, and the data obtained for both populations were compared. Sequencing data of several intermediate D12S391 alleles designated 17.3, 18.3, and 19.3 are also presented.     

Forensic and population genetic analysis of Serbian population using 21 STR loci of GlobalFiler™ PCR amplification kit

Autosomal short tandem repeats (STRs) have been widely used in forensic investigations. Prior to the application of any DNA based identification method, it is essential to estimate the allele frequencies and forensic statistical parameters of targeted STR loci in each population in order to provide a more precise reference database for forensic investigation. The GlobalFiler™ Kit is a multiplex assay that combines the 13 original CODIS loci with 7 non-overlapping loci from the expanded European Standard Set (ESS), as well as the highly discriminating SE33 locus, two Y-based loci and the sex determining maker, Amelogenin. The full complement of loci in the GlobalFiler™ Kit are: D13S317, D7S820, D5S818, CSF1PO, D1S1656, D12S391, D2S441, D10S1248, D18S51, FGA, D21S11, D8S1179, vWA, D16S539, TH01, D3S1358, AMEL, D2S1338, D19S433, DYS391, TPOX, D22S1045, SE33 and a Y-specific insertion/deletion locus (Yindel). The 6-dye GlobalFiler™ PCR Amplification kit (ThermoFisher Scientific) comprises 21 autosomal STRs have already been proven to be able to provide reliable DNA profiling results and enhance the power of discrimination between individuals. In this study, we are presenting an analysis of GlobalFiler STR loci on 209 unrelated individuals from Serbia.     

Validation and casework testing of the BioPlex-11 for STR typing of telogen hair roots

A new STR typing strategy has been developed allowing the simultaneous amplification and subsequent analysis of 11 polymorphic systems with amplicon sizes smaller than 270bp. The multiplex amplification reaction includes six STR loci from the European standard set of loci (ESS) for DNA databases (D3S1358, D8S1179, D21S11, THO1, FGA and VWA) as well as four additional STR systems selected for their robustness (D2S1338, D12S391, TPOX and D5S818) together with the sex-specific locus amelogenin. After PCR amplification, the multiplex reaction is splitted into two sets of STR multiplexes by using biotin labelled primers only for one set. Using streptavidin-coated Sepharose beads five STR systems are separated from the other six systems prior to being analysed in two different runs on a capillary gel electrophoresis instrument. The multiplex system was developed and tested especially for the use in forensic casework if only limited amounts or highly degraded DNA is available, for instance, when isolated from telogen hair roots.     

Genetic polymorphism of 14 non-CODIS STR loci for forensic use in southeast China population

We investigated 14 polymorphic STR loci (D1S2142, D2S1360, D3S1545, D7S1517, D10S2325, D12S391, D13S1492, D14S306, D15S659, D16S3253, D18S1270, D19S253, D20S470, D21S1437) which are not included in the standard sets of forensic loci (CODIS) in a sample of 216 unrelated healthy southeast Chinese individuals. The studied loci were highly informative and did not show departures from Hardy-Weinberg equilibrium. The accumulated powers of discrimination and power of exclusion for the 14 loci were 99.9999999999 and 99.999998%, respectively. No linkage was observed between the 14 loci and the traditional set of STR markers included in commercially available kits (the AmpFLSTR IdentifilerTM 15 System loci). We thus considered the studied 14 STRs are informative and when necessary, can be used as the candidate genetic markers in the study and application in genetics and forensic practice.     

温州汉族人群D12S391、D18S865基因座遗传多态性研究

目的研究短串联重复序列D12S391、D18S865的遗传多态性及法医学应用价值。方法应用聚合酶链反应(PCR)、聚丙烯酰胺凝胶电泳(PAGE)及银染显带技术对温州地区汉族454名无关个体的D12S391、D18S865位点进行分型。结果D12S391观察到11个等位基因,50种基因型;D18S865观察到7个等位基因,21种基因型。基因型频率的分布符合Hardy-Weinberg平衡,两个基因座的观测杂合度(H)分别为0.7974、0.7379;个体识别能力(DP)分别为0.9566、0.9077;多态信息含量(PIC)分别为0.8260、0.7279。结论两个STR基因座具有较高杂合度,等位基因分布符合Hardy-Weinberg平衡(P>0.5),在法医学应用和群体遗传学研究中较高的价值。     

Developmental validation of a single-tube amplification of the 13 CODIS STR loci, D2S1338, D19S433, and amelogenin: the AmpFlSTR Identifiler PCR Amplification Kit

Analysis of length polymorphism at short tandem repeat (STR) loci utilizing the polymerase chain reaction (PCR) process has proven to be an ideal assay for human identification purposes. The short length of STR loci coupled with the amplification of target sequence through PCR allows for a robust, sensitive, and specific assay for highly polymorphic markers. A multiplex containing fifteen STR loci plus the gender-determining locus Amelogenin was developed to provide a single amplification/detection of all CODIS (Combined DNA Index System) STR loci (CSF1PO, D3S1358, D5S818, D7S820, D8S1179, D13S317, D16S539, D18S51, D21S11, FGA, TH01, TPOX, and vWA) as well as two internationally-accepted STRs (D2S1338 and D19S433). By incorporating five-dye fragment analysis technology and non-nucleotide linkers, previously optimized AmpFlSTR kit primer sequences have been maintained. This kit has been developed in accordance with the standards of the forensic community as defined by the DNA Advisory Board. Validation studies were performed to include developmental validation, and the results support the use of the AmpFlSTR Identifiler PCR Amplification Kit for human identity and parentage testing.     

D6S1043和D12S391基因座在亲权鉴定中的应用

目的研究D6S1043和D12S391基因座在亲权关系鉴定案件中的应用价值。方法应用荧光标记复合扩增系统对日常检案中所收集的192名汉族无关个体血样DNA进行PCR扩增,用ABI3100-Avant遗传分析仪对扩增产物进行毛细管电泳,用GeneMapperv3.2软件进行基因分型,统计分析D6S1043和D12S391基因座的多态信息。结果在D6S1043和D12S391基因座分别发现12个等位基因,它们在中国汉族人群中的个体识别能力分别为0.9656和0.9510,二联体非父排除率分别为0.573和0.510,三联体非父排除率分别为0.731和0.679。结论D6S1043和D12S391基因座具有高度多态性,在亲权鉴定中具有重要应用价值。     

Practical applications of genotypic surveys for forensic STR testing

Legitimate genotype frequency estimation for multiallelic loci relies on component allele frequencies, as population surveys represent only a fraction of possible DNA profiles. Multilocus genotypes from two ethnic human populations, African American (n=195) and U.S. Caucasian (n=200), were compiled at 13 STR loci that are used worldwide in forensic investigation (D3S1358, vWA, FGA, D16S539, TH01, TPOX, CSF1PO, D8S1179, D21S11, D18S51, D5S818, D13S317, and D7S820). Sex-specific AmpFlSTR multiplexes provided stringent PCR-based STR typing specifically optimized for multicolor fluorescence detection. Heterozygosity at each STR locus ranged from 0.57 to 0.89 and encompassed from seven (TH01) to twenty-one (D21S11) alleles. Homozygosity tests, tests based on the distinct numbers of observed homozygous and heterozygous classes, log likelihood ratio tests, and exact tests assessed that the degree of divergence from theoretical Hardy-Weinberg proportions for all 13 STRs does not have practical consequence in genotype frequency estimation. Departures from linkage equilibrium, between loci, that imposed significance to forensic calculations were not indicated by observed variance of the number of heterozygous loci or Karlin interclass correlation tests. For forensic casework, reliable multilocus profile estimates may be obtained from the product of component genotype frequencies, each calculated through application of the Hardy-Weinberg equation to population database allele frequency estimates reported here. The average probability that two randomly selected, unrelated individuals possess an identical thirteen-locus DNA profile was one in 1.8x10(15) African Americans and one in 3.8x10(14) U.S. Caucasians.     

New alleles and mutational events at 14 STR loci from different German populations

The molecular origin of DNA mutations and the mutation rates were analyzed at 14 short tandem repeat (STR) loci with samples from trio cases derived from 10 different German population samples. STR loci comprised of D2S1360, D3S1744, D4S2366, D5S2500, D6S474, D7S1517, D8S1132, D10S2325, D12S391, D18S51, D19S246, D20S480, D21S226, and D22S689. In a total of 488 meioses, 16 isolated genetic inconsistencies in 8 different STRs were observed, whereas no mutations were found at the other loci. The data of five mutations suggested the presence of silent or null alleles due to sequence variation in primer binding site. This could be confirmed for four suspected cases by the use of alternative primer sets and by DNA sequence analyses. Furthermore, this study revealed nine new allelic variants at five different loci.     

Developmental validation of 15 X chromosomal short tandem repeat markers

The use of X chromosomal short tandem repeat (STR) markers has been greatly increasing in the forensic setting. Using guidelines set forth previously for the validation of autosomal and Y STRs, aspects of the feasibility of routine X chromosomal STR use were evaluated. Two mini-X chromosomal STR multiplexes capable of amplifying 15 total markers were developed and utilized to determine allele nomenclature, allele/genotype frequencies, mutation rates, and linkage between markers. Additionally, a concordance study between these multiplexes and a commercially available kit was performed. Here, the authors present an overview of this extensive developmental validation study.     

Effect of low-dose radiation on mutation rates of STR loci commonly used in forensic casework

The possible effect of low-dose radiation on STR markers in people exposed to radiation during their professional activities was estimated in this study. We evaluated mutation rates in 17 forensic STR loci typed by the COrDIS-18 kit (CSF1PO, D10S1248, D12S391, D13S317, D16S539, D18S51, D21S11, D2S441, D3S1358, D5S818, D7S820, D8S1179, FGA, SE33, TH01, TPOX, and vWA) in 78 families (father–mother–child) with one parent exposed to low-dose radiation before fertilization. Five mutations were observed. In two cases, the new alleles were delivered from the non-exposed parent. The calculated mutation rate for the 17 studied STR loci in families appeared to be in good concordance with data published for normal populations. No evidence for an elevated mutation rate in STR markers after low-dose radiation was found.     

4个YSTRs基因座的复合扩增

目的 建立DYS19、DYS389 Ⅰ、DYS389 Ⅱ、DYS385复合扩增体系。方法 遴选Y STRs基因座的引物，分别用FAM、TAMRA、TET标记DYS19、DYS385、DYS389 Ⅰ、DYS389 Ⅱ，优化扩增条件，考察扩增体系的个体识别能力、灵敏度、种属特异性及突变情况。结果 所建立的4基因座Y STRs复合扩增体系分型清晰，单倍型多样性达0.989，且特异性好，灵敏度高(1ng DNA)，未观察到突变。结论 所建立的4个Y STRs基因座复合扩增方法适合法医学应用。     

Population genetic data of 15 tetrameric short tandem repeats (STRs) in Berbers from Morocco

The allele frequency distribution of 15 short tandem repeats (STR) loci contained in the AmpFlSTR Identifiler PCR Amplification Kit (Applied Biosystems), was determined in two Berber populations from Asni and Bouhria, in Central and Eastern Morocco, respectively. A total of 209 individuals were typed. No deviations from the Hardy-Weinberg equilibrium were observed for Asni at the 15 STRs loci whereas for the Bouhria samples, two loci (D5S818 and TH01) showed significant departures from Hardy-Weinberg expectations (after Bonferroni's correction). All loci are highly polymorphic and population differentiation tests showed that the Moroccan samples from Asni and Bouhria have significant differences in 4 out of 15 loci (D21S11, D7S820, D16S539 and TPOX). The aim of the study was to obtain accurate allele frequencies relevant for forensic applications. Comparative analyses between our population data and other population samples gathered from the literature are also presented.     

STR data for the 15 loci from three minority populations in Guangxi municipality in South China

In this study, allele frequencies for the 15 STRs (D8S1179, D21S11, D7S820, CSF1PO, D3S1358, TH01, D13S317, D16S539, D2S1338, D19S433, vWA, TPOX, D18S51, D5S818, FGA) included in the AmpFLSTR Identifiler kit were obtained from total of 450 unrelated individuals belonging to three minority populations including Miao (n=141), Yi (n=165) and Hui (n=144) in Guangxi municipality, South China. GENEPOP v3.4 and PowerStats v1.2 was used to analyze the genotype data. This result indicates that the studied STR loci of these three minorities are useful in forensic testing and population genetics studies.     

Population distribution of six PCR-amplified loci in Madeira Archipelago (Portugal).

Frequency data of the short tandem repeat (STR) loci HUMTH01, HUMVWA31/A, HUMF13A1, HUMFES/FPS, D12S391 and HUMFIBRA/FGA were determined in blood stains obtained from a population of unrelated individuals from the Madeira Archipelago. The observed genotype distribution showed no significant deviation from the Hardy-Weinberg equilibrium and there was no evidence for association of alleles among the six loci. Population data showed a combined discrimination power of 0.9999998 and a chance of exclusion of 0.99597. The frequencies are similar to those of other compared caucasian populations but significant differences were found between the Madeira population and Japanese, Chinese, Greenland Eskimos and Quechua Amerindians. The six loci studied, together proved to be highly discriminating and valuable for forensic cases.     

Allele frequencies of 15 STRs in the Calchaqui Valleys population (North-Western Argentina)

Allele frequencies for 15 short tandem repeat (STR) loci were obtained from a sample of 110 individuals from the Calchaqui Valleys population (North-Western Argentina). The combined power of exclusion and combined power of discriminating for the 15 tested STR loci were 0.999964 and 0.9999999999999998, respectively. Matching probability was 1 in 4.58 × 10(15). Therefore, it may be concluded that the set of 15 STRs included in the AmpF STR Identifiler kit, represents a powerful tool for forensic applications, paternity testing and population genetics studies in the Calchaqui Valleys population.     

Genetic profiling of a central Venezuelan population using 15 STR markers that may be of forensic importance

The AmpFlSTR Identifiler kit has recently been accepted for use in DNA databasing of forensic samples in the FBI's National DNA Index System. In the present study, we used this kit to analyze the allele distribution of 15 short tandem repeat markers (STR) in individuals living in Caracas city, Venezuela. The allele frequencies of two of these STR, D2S1338 and D19S433, have not previously been reported for this or any other Latin American population. The results indicate that for the population here studied, the 15 STR tested are useful markers for paternity testing and forensic casework.