Sequence polymorphism of the mitochondrial DNA control region in the Slovenian population

The forensic application of mitochondrial DNA (mtDNA) typing requires large and regionally well-defined databases. To expand the database for forensic identification purposes in Slovenia, the mtDNA control region sequences of the hypervariable regions HVI and HVII were determined in a population of 129 maternally unrelated Slovenians, using a fluorescent-based capillary electrophoresis sequencing method. A total of 111 different haplotypes resulting from 124 polymorphic positions (80 polymorphic positions in HVI and 44 in HVII) were found. Of these, 101 mtDNA types were unique, 6 haplotypes were shared by 2 individuals, 1 haplotype by 3 individuals, 2 haplotypes by 4 individuals, and the most common haplotype was found in 5 individuals. The most frequent haplotypes in the Slovenian population ,263(G), 315.1(C) and 263(G), 309.1(C), 315.1(C) are also the most common in other European populations. The data support the concept that these haplotypes may represent a common European mtDNA sequence types. The sequence poymorphisms were compared to the databases of west Austria and central Italy and the HVI and HVII sequence matching probabilities within and between populations were calculated. It is 1.1–4.5 times more likely to find a sequence match in a random pair of Slovenians than in a random Slovenian-Italian pair and in a random Slovenian-Austrian pair. The length heteroplasmy in the homopolymeric C-stretch regions located at nucleotide positions 16184–16193 in HVI and at positions 303–315 in HVII was observed in 17% and 8% of individuals, respectively. A statistical estimate of the results for this population showed the random match probability and the genetic diversity of 1.16% and 0.996, respectively.Electronic Supplementary Material Supplementary material is available in the online version of this article at http://dx.doi.org/10.1007/s00414-003-0394-3     

Sequence polymorphisms of the mitochondrial DNA control region and phylogenetic analysis of mtDNA lineages in the Japanese population

Sequence polymorphisms of the hypervariable mitochondrial DNA (mtDNA) regions HVI and HVII, and coding region polymorphisms were investigated in 211 unrelated individuals from the Japanese population. Sequence comparison of the HVI and HVII regions led to the identification of 169 mitochondrial haplotypes defined by 147 variable positions. Among them 145 types were observed in only 1 individual; the other 24 types were shared by 2 or more individuals. The gene diversity was estimated at 0.9961, and the probability of two randomly selected individuals from the population having identical mtDNA types was 0.86%. We also established phylogenetic haplogroups in the Japanese population based on the coding and control region polymorphisms and compared the haplotypes with those in other Japanese, Korean and Chinese populations. As a result, three new subhaplogroups, G4a, G4b, and N9b, and several haplotypes specific for the Japanese and Korean populations were identified. The present database can be used not only for personal identification but also as an aid for geographic or phenotype (race) estimation in forensic casework in Japan.Electronic Supplementary Material Supplementary material is available for this article if you access the article at     

Population study and evaluation of 20 Y-chromosome STR loci in Germans

The nine European minimal haplotype (EMH) loci, the two SWGDAM loci and five further single-copy Y-chromosomal short tandem repeats (Y-STRs) DYS446, DYS447, DYS448, DYS449, DYS463, and the multicopy loci DYS464 were evaluated in the German population groups Dresden, Hamburg, Rostock, Munich, and the Sorbs who are a Slavic-speaking minority in Lusatia. Highest gene diversities in all populations were shown for DYS464, DYS385, and DYS449 (D=0.8559–0.9486). The haplotype diversity for the European minimal haplotype loci ranged between 0.9852 for Sorbs and 0.9983 for the Hamburg population showing that there is a significant portion of haplotypes which could not be resolved. Advanced typing using DYS446, DYS447, DYS448, DYS449, DYS463, and DYS464 discriminated all non-related individuals of the Dresden, Hamburg, and Rostock populations. Evaluation of the Y-STRs was accomplished by sequence analysis of all allelic fragments of the allelic ladders and microvariant alleles of DYS385 and the determination of the amounts of stutter products of the loci. Electronic Supplementary Material Supplementary material is available for this article at     

Mitochondrial DNA D-loop hypervariable regions: Czech population data

In order to identify polymorphic sites and to find out their frequencies and the frequency of haplotypes, the complete D-loop of mitochondrial DNA (mtDNA) from 93 unrelated Czech Caucasians was sequenced. Sequence comparison showed that 85 haplotypes were found and of these 78 were unique, 6 were observed twice and 1 was observed three times. Genetic diversity (GD) was estimated at 0.999 and the probability of two randomly selected sequences matching (random match probability, RMP) at 1.2%. Additionally these calculations were carried out for hypervariable regions 1, 2 (HV1, HV2), for the area between HV1 and HV2 and for the area of the hypervariable region HV3. The average number of nucleotide differences (ANND) was established to be 10.2 for the complete D-loop. The majority of sequence variations were substitutions, particularly transitions. Deletions were found only in the region where HV3 is situated and insertions in the same place and in poly-C tracts between positions 303 and 315 in HV2. A high degree of length heteroplasmy was found especially in the regions of poly-C tracts between positions 16184 and 16193 in HV1 and between positions 303 and 315 in HV2. Position heteroplasmies were found in two cases.Electronic Supplementary Material Supplementary material is available in the online version of this article at http://dx.doi.org/10.1007/s00414-003-0407-2     

Sequence analysis of mitochondrial DNA hypervariable region I in Thai individuals

Nucleotide sequence analysis of hypervariable region I (HVRI) in human mitochondrial DNA (mtDNA) was investigated in 100 unrelated Thai individuals. A total of 85 variable sites and 423 base substitutions, which consisted of 390 nucleotide transitions and 33 nucleotide transversions were found. The following nucleotide substitutions were found: 48% at 16,223, 31% at 16,304, 30% at 16,332, and 26% at 16,129, respectively. Transition from T to C (43.7%) was the most frequent substitution. The nucleotide insertions were found at two sites with T at position 16,188 and C at position 16,194. Eighty-two haplotypes were investigated of which 72 haplotypes were unique. The most frequent haplotypes (16,108T-16,129A-16,162G-16,172C-16,304C and 16,260T-16,298C-16,355T-16,362C) were observed. From position 16,180 to 16,193, thirteen patterns of polycytosine or C-stretch were observed, whereas 68 Thai individuals were found to be similar to the references. The genetic diversity, random match probability, and discrimination power were estimated to be 0.9943, 0.0156, and 0.9844, respectively.     

Mitochondrial DNA variation in the Sindh population of Pakistan

The present study was undertaken to investigate the control region of mitochondrial DNA for forensic discrimination and to explore the ethno-linguistic affiliations among ethnic groups of Sindh province, Pakistan. A total of 115 individuals, from six major ethnic/isonym groups, namely, Bijarani, Chandio, Ghallu, Khoso, Nasrani and Solangi, have been studied. We identified 88 haplotypes, defined by the particular set of nucleotides; of these, 70 haplotypes were unique in the investigated population. In addition, 82% sequences were observed once, 12% twice and 5.2% thrice. The most common South Asian haplogroup in six ethnic groups of Sindh, are; M (42%) and R (6.9%), whereas West Eurasian haplogroups were N (6.9%), W (6.9%), J (1.7%), U (23.4%), H (9.5%) and T (0.86%). A random match probability between two unrelated individuals was found to be 0.06%, while genetic diversity varied from 0.991 to 0.998. The high nucleotide diversity and the low random match probability of the mtDNA control region make it a beneficial tool for forensic discrimination as well as useful to the evolutionary biologist. This work is an important contribution towards establishing a National Mitochondrial DNA Database in Pakistan. Currently, the Human Genetic department at the University of Health Sciences Lahore has the depository.     

Hungarian mtDNA population databases from Budapest and the Baranya county Roma

To facilitate forensic mtDNA testing in Hungary, we have generated control region databases for two Hungarian populations: 211 individuals were sampled from the urban Budapest population and 208 individuals were sampled from a Romani (“gypsy”) population in Baranya county. Sequences were generated using a highly redundant approach to minimize potential database errors. The Budapest population had high sequence diversity with 180 lineages, 183 polymorphic positions, and a random match probability of 1%. In contrast, the Romani population exhibited low sequence diversity, with only 56 lineages, 109 segregating sites, and a random match probability of 8.8%. The mtDNA haplogroup compositions of the two populations were also distinct, with the large proportion of haplogroup M samples (35%) in the Roma the most obvious difference between the two populations. These factors highlight the importance of considering population structure when generating reference databases for forensic testing purposes. Comparisons between our Romani population sample and other published data indicate the need for heightened caution when sampling and using mtDNA databases of small endogamous populations. The Romani populations that we compared showed significant departures from genetic uniformity. Electronic supplementary material Supplementary material is available in the online version of this article at and is accessible for authorized users.     

Human Y-chromosomal STR haplotypes in a Kurdish population sample

In an Iraqi Kurdish population sample (n = 101), seven polymorphic STR loci of the Y-chromosome (DYS19, 389, 390, 391, 392, 393, and DXYS156-Y) were typed, with DYS389 being subtyped for its four segments. The haplotype diversity was 97.83% and 82 different haplotypes were observed. The Kurds shared some Y-types with neighbouring south Turks but strikingly few with Germans: it is 20–30 times more likely to find a sequence match in a random pair of Kurds than in a random Kurd-German pair. Received: 25 September 1998 / Received in revised form: 4 November 1998 / Accepted: 5 November 1998     

Mitochondrial DNA sequences for 118 individuals from northeastern Spain

A population database was generated from 118 unrelated Caucasoid individuals living in Spain. Sequence polymorphisms of the mitochondrial DNA (mtDNA) control region, hypervariable regions I and II (HVRI and HVRII) were determined using the polymerase chain reaction (PCR) and direct sequencing. A total of 102 different sequences were found as defined by 105 variable positions. The most common sequence occurred six times, and this sequence is also the most frequent in other European populations such as Austria, Germany and Britain. The mean pair-wise difference for the two HVR regions taken together was 7.74. The study revealed that transitions made up the majority of the variations (88%), whereas we observed a significantly lower frequency of transversions (8%). Also one individual in this study was observed with two positions of heteroplasmy at nucleotides 150 (C/T) and 153 (G/A). A statistical estimate of the results for this population showed a genetic diversity of 0.99. The probability of two random individuals showing identical mtDNA haplotypes is 1.3%. In order to use the mtDNA analysis in forensic casework, we consider that it is of crucial importance to know the frequency of the different sequences of mtDNA, and this data base study could be a useful tool to statistically evaluate the results. Received: 12 July 1999 / Accepted: 11 April 2000     

Y chromosome STR allelic and haplotype diversity in a Rwanda population from East Central Africa

We have analyzed 17 Y-chromosomal STR loci in a population sample of 69 unrelated male individuals of the Rwanda-Hutu population from East Central Africa using an AmpFlSTR® Yfiler? PCR amplification kit. A total of 62 unique haplotypes were identified among the 69 individuals studied. The haplotype diversity was found to be 0.9970 for this population. The gene diversity ranged from 0.1130 (DYS392) to 0.7722 (DYS385). Comparison of populations in this study with twenty-five other national and global populations using Principal Co-ordinate Analysis (PCA) and phylogenetic molecular analysis using a genetic distance matrix indicates a delineation of all the African populations from other unrelated populations. The results of population pair-wise Fst p values indicate statistically significant differentiation of the Rwandan population when compared with 25 other global populations including four African populations (p = 0.0000). Analysis of Molecular Variance (AMOVA) of the Rwanda population with four other African populations indicated a 93% variance within populations and 7% variance among the five populations. A data base search of the 62 haplotypes yielded only one non-African haplotype match, suggesting these haplotypes are unique to the African continent.     

Polish population study on Y chromosome haplotypes defined by 18 STR loci

Polymorphism of 18 STR loci specific to the human Y chromosome (DYS19, DYS388, DYS389I, DYS389II, DYS390, DYS391, DYS392, DYS393, DYS426, DYS437, DYS438, DYS439, DYS460, GATA H4.1, DYS385 a/b, and YCAII a/b) was evaluated by means of a multiplex (octadecaplex) PCR reaction and capillary electrophoresis in a Polish population sample of 208 unrelated males. A total of 192 different haplotypes and 183 unique haplotypes were identified. The observed haplotype diversity was 0.998, while discrimination capacity was 92.3%. DYS389 was shown to be the most valuable in discrimination of similar haplotypes, whereas DYS388, DYS393, DYS426, and DYS438 did not affect the discrimination power of the multiplex.Electronic Supplementary Material Supplementary material is available for this article at .     

Predicting sampling saturation of mtDNA haplotypes: an application to an enlarged Portuguese database

An enlarged mtDNA database (n=549) for the Portuguese population, comprising HVRI and HVRII regions is reported. This database was used to test the effect of sample size on the estimation of relevant parameters such as haplotype diversity, number of different haplotypes, nucleotide diversity and number of polymorphic positions. Simulations were performed generating sets of random subsamples of variable sizes (n=50, 100, 200, 300 and 400). The results show that while haplotype and nucleotide diversities do not vary significantly with sample size, the numbers of haplotypes and polymorphic positions rise continuously inside the tested interval. These trends are interpretable by the evolution of the proportions of sequences that are found once or twice, which drop dramatically as sample size increases, with the corresponding rise in the frequency of those encountered 3 times or more. The generated data were also used to extrapolate saturation curves for the referred parameters. When considering for instance the number of haplotypes, it is shown that a sample size of 1,000 individuals is required for practical saturation (defined as the point where a sample size increase of 100 individuals corresponds to an increment in the diversity measure below 5%). For HVRII the same level is reached at n=900 and n=1,300 is needed when both regions are analysed simultaneously. Consequently, we can infer that currently used sample sizes are still rather inadequate for both anthropological and forensic purposes.Electronic Supplementary Material Supplementary material is available in the online version of this article at http://dx.doi.org/10.1007/s00414-003-0424-1     

Mutations in 14 Y-STR loci among Japanese father-son haplotypes

In the present study 161 Japanese father/son haplotype transfers in 147 pedigrees were analyzed at 14 Y-STRs with two multiplex PCR-based typing systems. Five isolated single repeat mutations were identified at the DYS389I, DYS439, Y-GATA-H4, DYS389II and DYS391 loci, and a pedigree showing triple alleles at the DYS385 locus (a duplicate locus) without allelic discrepancy between the father and son was also observed. The overall mutation rate estimated across the 14 Y-STRs in the Japanese population was 0.22%/locus/meiosis (95% C.I. 0.09–0.51%). This rate was not significantly different (p>0.05) from those of autosomal STRs and Y-STRs in other populations, including German, Austrian, Polish and Norwegian populations. Furthermore, 138 haplotypes were identified in 147 pedigrees with a haplotype diversity value of 0.9983. Therefore, a combination of the two systems should permit effective analysis with sufficient discriminatory power.     

Global population variability in Promega PowerPlex CS7, D6S1043, and Penta B STRs

Supplementary short tandem repeats (STRs) can be added to forensic DNA analyses when core markers fail to provide sufficient discrimination power in identity and relationship testing. We combined D6S1043 and Penta B with Promega's PowerPlex CS7 supplementary STR kit, comprising Pentas D and E plus LPL, F13A01, FES/FPS, F13B, and Penta C. The nine STRs were typed in 941 individuals from 51 diverse populations of the CEPH Human Genome Diversity Panel (HGDP-CEPH), and we report allele frequency estimates plus rare alleles identified. Both Penta B and D6S1043 show highly informative variation in all populations, exceeding most CS7 STRs and raising cumulative random match probabilities by at least two orders of magnitude. However, Penta B genotype distributions show an excess of homozygotes across all HGDP-CEPH population groups indicating likely allele dropout from uncharted SNP or Indel variation at the primer sites chosen to type this STR. The first sequence analysis of common regular and rare intermediate D6S1043 alleles is reported. D6S1043 .3 intermediate alleles were found to occur at a high frequency in Native Americans, providing scope for differentiation of this group.     

Polymorphism of the mitochondrial DNA control region in Italians

Mitochondrial DNA sequences of the hypervariable regions HVI and HVII were analysed in 83 Caucasians living in central Italy to expand the database for forensic identification purposes, and 75 different haplotypes resulting from 62 polymorphic positions in HVI and 44 in HVII were observed. The most frequent haplotype (263G, 309.1C, 315.1C) was shared by 7 individuals, 2 haplotypes were shared by 2 individuals, and 72 were unique. The genetic diversity was found to be 0.99 and the random match probability 1.9%. A condition of sequence heteroplasmy was found in only one case at nt 16311, whereas a length heteroplasmy was found in the homopolymeric stretch of cytosines 303–315. Our results indicate that in direct sequencing beyond the poly-cytosine stretch, the overlap is due to length heteroplasmy, whereas the blurred signal occurs when the stretch is composed of more than 10 cytosines. Received: 14 February 2000 / Accepted: 23 May 2000     

Population data for 17 Y-chromosome STRs in a sample from Apulia (Southern Italy)

The 17 Y-STR loci included in the AmpFLSTR Yfiler PCR Amplification Kit were analyzed in 98 unrelated healthy males from Apulia (Southern Italy). A total of 97 different haplotypes were identified, of which 96 haplotypes were unique and 1 occurred twice. Allele frequencies for each Y-STR locus in pooled sample and estimated value of gene diversity (GD) were evaluated. The lowest value of GD was observed for DYS392 (0.126) and the highest one (0.936) for DYS385. The HD (haplotype diversity) for the studied Y-STR set showed a value of 0.9994, with an HMP (haplotype match probability) value of 0.0006, while the overall DC was 98.98%. Microvariant alleles were found for the DYS458 and DYS385 markers and sequenced.Furthermore, Φ_st-based genetic distance computation and pair-wise analysis of molecular variance (AMOVA) test were carried out. When comparing our population with the Apulia sample previously investigated, the AMOVA analysis detected no evidence for significant differentiation. The comparison with all Italian populations submitted to the YHRD website showed no relevant differences with all Southern Italian populations (San Giorgio La Molara, Belvedere, Trapani and Catania) and significant genetic deviation with all Northern Italian populations (Udine, Biella, La Spezia, Modena, Ravenna, Marche and North Sardinia). Moreover, the other populations and meta-populations belonging to the whole Mediterranean area (Croatia, Macedonia, Albania, Greece, Turkey, Israel, Libya, Tunisia, Algeria, Morocco and Spain) were different from our Apulia sample. The data were submitted to YHRD.     

Sequence polymorphism of the mitochondrial DNA hypervariable regions I and II in 205 Singapore Malays

Mitochondrial DNA sequences of the hypervariable regions HV1 and HV2 were analyzed in 205 unrelated ethnic Malays residing in Singapore as an initial effort to generate a database for forensic identification purposes. Sequence polymorphism was detected using PCR and direct sequencing analysis. A total of 152 haplotypes was found containing 152 polymorphisms. Out of the 152 haplotypes, 115 were observed only once and 37 types were seen in multiple individuals. The most common haplotype (16223T, 16295T, 16362C, 73G, 146C, 199C, 263G, and 315.1C) was shared by 7 (3.41%) individuals, two haplotypes were shared by 4 individuals, seven haplotypes were shared by 3 individuals, and 27 haplotypes by 2 individuals. Haplotype diversity and random match probability were estimated to be 0.9961% and 0.87%, respectively.     

HVI and HVII mitochondrial DNA data in Apaches and Navajos

Most mtDNA studies on Native Americans have concentrated on hypervariable region I (HVI) sequence data. Mitochondrial DNA haplotype data from hypervariable regions I and II (HVI and HVII) have been compiled from Apaches (N=180) and Navajos (N=146). The inclusion of HVII data increases the amount of information that can be obtained from low diversity population groups. Less mtDNA variation was observed in the Apaches and Navajos than in major population groups. The majority of the mtDNA sequences were observed more than once; only 17.8% (32/180) of the Apache sequences and 25.8% of the Navajo sequences were observed once. Most of the haplotypes in Apaches and Navajos fall into the A and B haplogroups. Although a limited number of haplogroups were observed, both sample populations exhibit sufficient variation for forensic mtDNA typing. Genetic diversity was 0.930 in the Apache sample and 0.963 in the Navajo sample. The random match probability was 7.48% in the Apache sample and 4.40% in the Navajo sample. The average number of nucleotide differences between individuals in a database is 9.0 in the Navajo sample and 7.7 in the Apache sample. The data demonstrate that mtDNA sequencing can be informative in forensic cases where Native American population data are used.     

Mitochondrial DNA analysis of Swedish population samples

As a contribution to the geographic coverage of EMPOP, currently the best available forensic mitochondrial DNA (mtDNA) database, a total of 299 Swedish individuals were analysed by sequencing of the first and second hypervariable regions of the mtDNA genome. In this sample set, a total of 179 different haplotypes were detected. The genetic diversity was estimated to be 0.9895 (±0.0023), and the random match probability was 1.39 %. The most abundant haplogroups were HV (including its subhaplogroups H and V) with a frequency of 46.5 %, followed by haplogroup U (including its subhaplogroup K) at 27.8 %, haplogroup T at 10.0 % and haplogroup J at 7.0 %, a distribution that is consistent with previous observations in other European populations.     

Mitochondrial DNA control region sequences in Koreans: identification of useful variable sites and phylogenetic analysis for mtDNA data quality control

We have established a high-quality mtDNA control region sequence database for Koreans. To identify polymorphic sites and to determine their frequencies and haplotype frequencies, the complete mtDNA control region was sequenced in 593 Koreans, and major length variants of poly-cytosine tracts in HV2 and HV3 were determined in length heteroplasmic individuals by PCR analysis using fluorescence-labeled primers. Sequence comparison showed that 494 haplotypes defined by 285 variable sites were found when the major poly-cytosine tract genotypes were considered in distinguishing haplotypes, whereas 441 haplotypes were found when the poly-cytosine tracts were ignored. Statistical parameters indicated that analysis of partial mtDNA control region which encompasses the extended regions of HV1 and HV2, CA dinucleotide repeats in HV3 and nucleotide position 16497, 16519, 456, 489 and 499 (HV1ex+HV2ex+HV3CA+5SNPs) and the analysis of another partial mtDNA control region including extended regions of HV1 and HV2, HV3 region and nucleotide position 16497 and 16519 (HV1ex+HV2ex+HV3+2SNPs) can be used as efficient alternatives for the analysis of the entire mtDNA control region in Koreans. Also, we collated the basic informative SNPs, suggested the important mutation motifs for the assignment of East Asian haplogroups, and classified 592 Korean mtDNAs (99.8%) into various East Asian haplogroups or sub-haplogroups. Haplogroup-directed database comparisons confirmed the absence of any major systematic errors in our data, e.g., a mix-up of site designations, base shifts or mistypings. Electronic supplementary material Supplementary material is available for this article at and accessible for authorised users.