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.     

Sequence polymorphisms of mtDNA HV1, HV2, and HV3 regions in the Malay population of Peninsular Malaysia

The uniparentally inherited mitochondrial DNA (mtDNA) is in the limelight for the past two decades, in studies relating to demographic history of mankind and in forensic kinship testing. In this study, human mtDNA hypervariable segments 1, 2, and 3 (HV1, HV2, and HV3) were analyzed in 248 unrelated Malay individuals in Peninsular Malaysia. Combined analyses of HV1, HV2, and HV3 revealed a total of 180 mtDNA haplotypes with 149 unique haplotypes and 31 haplotypes occurring in more than one individual. The genetic diversity was estimated to be 99.47%, and the probability of any two individuals sharing the same mtDNA haplotype was 0.93%. The most frequent mtDNA haplotype (73, 146, 150, 195, 263, 315.1C, 16140, 16182C, 16183C, 16189, 16217, 16274, and 16335) was shared by 11 (4.44%) individuals. The nucleotide diversity and mean of pair-wise differences were found to be 0.036063 ± 0.020101 and 12.544022 ± 6.230486, respectively.     

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     

Sequence polymorphism of mitochondrial DNA in Japanese individuals from Gifu Prefecture

Sequence polymorphisms of the hypervariable region HV1 in mitochondrial DNA (mtDNA) were analyzed in a sample of 137 unrelated Japanese individuals living in Gifu Prefecture (central region of Japan) using polymerase chain reaction amplification and direct sequencing. Eighty-two different haplotypes resulting from 81 variable sites were found in the mtDNA HV1 region between positions 16061 and 16450. The most frequent haplotype (16223T, 16362C) was shared by ten individuals. The genetic diversity and the genetic identity were 0.985 and 0.022, respectively. The C-stretch region located around position 16189 was observed in 23.4% of this population sample. Sequence heteroplasmy at the position 16103 (A/G) was found in one individual.     

Haplotype diversity in mitochondrial DNA hypervariable region in a population of southeastern Brazil

Brazilian population derives from Native Amerindians, Europeans, and Africans. Southeastern Brazil is the most populous region of the country. The present study intended to characterize the maternal genetic ancestry of 290 individuals from southeastern (Brazil) population. Thus, we made the sequencing of the three hypervariable regions (HV1, HV2, and HV3) of the mitochondrial DNA (mtDNA). The statistical analyses were made using Arlequin software, and the median-joining haplotype networks were generated using Network software. The analysis of three hypervariable regios showed 230 (79.3 %) unique haplotypes and the most common haplotype was “263G” carried by 12 (4.1 %) individuals. The strikingly high variability generated by intense gene flow is mirrored in a high sequence diversity (0.9966?±?0.0010), and the probability of two random individuals showing identical mtDNA haplotypes were 0.0068. The analysis of haplogroup distribution revealed that 36.9 % (n?=?107) presented Amerindian haplogroups, 35.2 % (n?=?102) presented African haplogroups, 27.6 % (n?=?80) presented European haplogroups, and one (0.3 %) individual presented East Asian haplogroup, evidencing that the southeastern population is extremely heterogeneous and the coexistence of matrilineal lineages with three different phylogeographic origins. The genetic diversity found in the mtDNA control region in the southeastern Brazilian population reinforces the importance of increased national database in order to be important and informative in forensic cases.     

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 polymorphism of the mitochondrial DNA control region in the population of Vojvodina Province,Serbia

In order to generate and establish the database for forensic identification purposes in Vojvodina Province (Serbia), the sequence of the hypervariable regions 1 (HV1) and 2 (HV2) of the mtDNA control region were determined in a population of 104 unrelated individuals from Vojvodina Province, using a fluorescent-based capillary electrophoresis sequencing method. A total of 93 different haplotypes were found, of these 83 mtDNA types were unique, nine haplotypes were shared by two individuals and one haplotype by three individuals. The variation of mtDNA HV1 and HV2 regions was confined to 116 nucleotide positions, of which 72 were observed in the HV1 and 44 in the HV2. A statistical estimate of the results for this population showed the genetic diversity of 0.9977 and the random match probability of 1.18%. Haplogroup H was the most common haplogroup (43.3%). Haplogroups observed at intermediate levels included clusters U (13.5%), T (10.6%), J (8.6%) and W (5.8%).     

Mitochondrial DNA extraction and typing from isolated dentin-experimental evaluation in a Korean population

This study reports mtDNA polymorphisms in both hypervariable segments HV1 and HV2 of the non coding D-loop region from 60 unrelated Koreans. In contrast to two previous Korean data base studies, mtDNA was extracted separately from pulp tissue and root dentin of teeth obtained from dentists. Dentin turned out to be a reliable source of mitochondrial DNA. This can be of practical importance in forensic identification case work after a long post-mortem interval since pulp decomposes rapidly. The extraction method is explained in detail. The mtDNA polymorphisms obtained from 60 teeth of unrelated Koreans were compared with the already existing Korean data base. Received: 27 October 1997 / Received in revised form: 2 Feburary 1998     

A database of mitochondrial DNA hypervariable regions I and II sequences of individuals from Slovakia

In order to identify polymorphic positions and to determine their frequencies and the frequency of haplotypes in the human mitochondrial control region, two hypervariable regions (HV1 and HV2) of the mitochondrial DNA (mtDNA) of 374 unrelated individuals from Slovakia were amplified and sequenced. Sequence comparison led to the identification of 284 mitochondrial lineages as defined by 163 variable sites. Genetic diversity (GD) was estimated at 0.997 and the probability of two randomly selected individuals from population having identical mtDNA types (random match probability, RMP) for the both regions is 0.60%.     

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.     

Mitochondrial DNA population data of HV1 and HV2 sequences from Japanese individuals

Mitochondrial DNA sequences of the hypervariable regions HV1 and HV2 were determined for 1204 unrelated Japanese individuals. A total of 741 different mtDNA haplotypes were found, 157 of which were seen in multiple individuals. Twenty-seven of these individuals showed point heteroplasmy. The most frequent haplotype (16223T-16362C-73G-263G-315.1C) was found in 31 individuals and the second most frequent haplotypes (16129A-16223T- 16362C-73G-152C-263G-309.1C-315.1C) was found in 24 individuals. The haplotypes diversity and random match probability were calculated to be 0.9969 and 0.0040, respectively.     

Expanding the forensic German mitochondrial DNA control region database: genetic diversity as a function of sample size and microgeography

Mitochondrial DNA control region sequences were determined in 109 unrelated German Caucasoid individuals from north west Germany for both hypervariable regions 1 (HV1) and 2 (HV2) and 100 polymorphic nucleotide positions (nps) were found, 63 in HV1 and 37 in HV2. A total of 100 different mtDNA lineages was revealed, of which 7 were shared by 2 individuals and 1 by 3 individuals. The probability of drawing a HV1 sequence match within the north west Germans or within published sets of south Germans and west Austrians is similar (within a factor of 2) to drawing a sequence match between any two of these three population samples. Furthermore, HV1 sequences of 700 male inhabitants of one village in Lower Saxony were generated and these showed a nearly linear increase of the number of different haplotypes with increasing number of individuals, demonstrating that the commonly used haplotype diversity measure (Nei 1987) for population samples tends to underestimate mtDNA diversity in the actual population. Received: 14 January 1999 / Accepted: 15 February 1999     

The results of an mtDNA study of 1200 inhabitants of a German village in comparison to other Caucasian databases and its relevance for forensic casework

Mitochondrial DNA control region sequences were determined in 1200 male volunteers from one village area of Lower Saxony for the hypervariable region 1 (HV1). The 154 variable positions found resulted in 460 different haplotypes with a haplotype diversity value of 0.98165. The number of different haplotypes showed a nearly linear increase with the number of individuals typed. The haplotype diversity approached saturation level at a value of approximately 0.981 after typing 400 individuals. Furthermore, the number of different haplotypes and the haplotype diversity were calculated for four short amplicons of HV1 in order to establish the most variable section with a high efficiency for forensic casework. Received: 29 January 2000 / Accepted: 24 April 2000     

A new database of mitochondrial DNA hypervariable regions I and II sequences from 162 Japanese individuals

A database of mitochondrial DNA (mtDNA) hypervariable region 1 (HV1) and region 2 (HV2) sequences of the mtDNA control region was established from 162 unrelated Japanese individuals. The random match probability and the genetic diversity for this database were 0.96% and 0.997, respectively. Length heteroplasmy in the C-stretch regions located around position 16189 in HV1 and 310 in HV2 was observed in 37% and 38% of the samples, respectively. A strategy using internal sequencing primers was devised to obtain confirmed sequences in these length heteroplasmic individuals. This database, combined with other mtDNA sequence databases from the Japanese population, will permit the significance of mtDNA match results to be properly reported in mtDNA typing casework in Japan.Received: 10 August 2000 / Accepted: 7 January 2001An erratum to this article can be found at     

Location and frequency of polymorphic positions in the mtDNA control region of individuals from Germany

In order to identify polymorphic positions and to determine their frequency in the human mitochondrial D-loop containing region, the mitochondrial DNA (mtDNA) control region of 200 unrelated individuals from Germany were amplified and directly sequenced. Sequence comparison led to the identification of 190 mitochondrial lineages as defined by 202 variable positions. The most frequently occurring lineage comprised 5 individuals, whereas 186 types of D-loop sequences were observed in only one individual. Of the sequences studied 7% are not unique but show at least one counterpart with an identical haplotype. The majority (61%) of the control regions investigated showed between four and eight nucleotide positions deviating from the reference sequence. The maximum number of deviations observed in a single control region was 18. The majority of the variable positions in the D-loop region (88%) are located within three hypervariable regions. Sequence variations are caused by nucleotide substitutions, insertions or deletions. As compared to insertions and deletions, nucleotide substitutions make up the vast majority of the mutations (90%). We have predominantly found transitions (75%) and a significantly lower frequency of transversions (15%) whereas insertions (6%) as well as deletions (4%) are rather rare. Upon sequencing the mitochondrial control region from 200 German Caucasians the genetic diversity was estimated at 0.99. The probability of two randomly selected individuals from a population having identical mtDNA types is 0.6%. Received: 21 May 1997 / Received in revised form: 9 September 1997     

Italian mitochondrial DNA database: results of a collaborative exercise and proficiency testing

This work is a review of a collaborative exercise on mtDNA analysis undertaken by the Italian working group (Ge.F.I.). A total of 593 samples from 11 forensic genetic laboratories were subjected to hypervariable region (HVS-I/HVS-II) sequence analysis. The raw lane data were sent to MtDNA Population Database (EMPOP) for an independent evaluation. For the inclusion of data for the Italian database, quality assurance procedures were applied to the control region profiles. Only eight laboratories with a final population sample of 395 subjects passed the quality conformance test. Control region haplogroup (hg) assignments were confirmed by restriction fragment length polymorphism (RFLP) typing of the most common European hg-diagnostic sites. A total of 306 unique haplotypes derived from the combined analysis of control and coding region polymorphisms were found; the most common haplotype--CRS, 263, 309.1C, 315.1C/ not7025 AluI--was shared by 20 subjects. The majority of mtDNAs detected in the Italian population fell into the most common west Eurasian hgs: R0a (0.76%), HV (4.81%), H (38.99%), HV0 (3.55%), J (7.85%), T (13.42%), U (11.65%), K (10.13%), I (1.52%), X (2.78%), and W (1.01%).     

Mitochondrial DNA in human hair shafts – existence of intra-individual differences?

The sequences of the hypervariable region 1 (HV1) of the mitochondrial DNA control region from multiple hair shafts from 10 unrelated individuals were compared to determine the frequency of differences in hairs from one individual. The extraction method described herein showed an average success rate of 67% for all 150 hair shafts tested in HV1. The mtDNA sequences from the hair shafts matched the sequences from the corresponding blood and saliva samples taken from the same donor and no evidence of heteroplasmy was found. The results emphasize the reliability of DNA extraction and mtDNA typing from human hair shafts for forensic purposes. Received: 16 July 1998 / Received in revised form: 31 August 1998     

Subtyping mtDNA haplogroup H by SNaPshot minisequencing and its application in forensic individual identification

Sequence variation of the hypervariable segments (HVS) I/II of mitochondrial DNA (mtDNA) and the haplogroup affiliation were determined in a sample of 271 Italian subjects. This analysis showed that 42% of the individuals could be ascribed to H, the most frequent haplogroup in European Caucasian populations. This fraction was then screened for specific single nucleotide polymorphisms located in the coding region to identify H subclades H1–H15. We set up two multiplex polymerase chain reactions and specific SNaPshot assays to investigate the frequency distribution of these subgroups in our population sample and to examine their usefulness in discriminating among commonly shared HVS I/II sequences. This allowed the assignment of a large portion of the mtDNAs (∼70%) to specific subhaplogroups, with H1 and H5 being the most represented. About two-thirds of the individuals sharing common HVS I/II sequences were subdivided and ascribed to specific H subhaplogroups with a significant reduction of the frequencies of the most common mtDNA haplotypes. Haplogroup H subtyping could thus be extremely useful in forensic identification when many samples have to be analysed and compared, avoiding excessive time-consuming and labor-intensive sequencing analysis.     

Phylogenetic classification of Japanese mtDNA assisted by complete mitochondrial DNA sequences

We investigated control and coding region polymorphisms in mitochondrial DNA (mtDNA) in 100 unrelated individuals from a Japanese population and determined the basal phylogenetic haplogroup lineages in all samples under updated information. Many of the basal phylogenetic haplogroup lineages assigned on East Asian mtDNA haplogroups corresponded to those previously established. However, new haplogroup lineages such as M7a2a, M7a2b, M7a2*, M7c1b, M11b2*, G2b*, D4c1b1a, D4g2b, A4*, A9, N9b*, B4d1, B4d2, and F1e were identified and established by complete sequencing. Although sequence comparison of the 1.15-kb control region identified 84 mitochondrial haplotypes, examination of coding region polymorphisms increased the total number of haplotypes to 91. Determination of the basal haplogroup lineages increased the discrimination power of mtDNA polymorphisms for personal identification and their usefulness in determining geographic origin in forensic casework in Japanese and other East Asian populations.     

Genetic analysis of human remains found in two eighteenth century Yakut graves at At-Dabaan

We extracted DNA from three skeletons belonging to the Yakut population, which were excavated from the At-Dabaan site (dating back 300 years) in the Sakha Republic (Russia). Ancient DNA was analyzed by autosomal STRs (short tandem repeats) and by the sequencing of the hypervariable region 1 (HV1) of the mitochondrial DNA (mtDNA) control region. The results showed that these three skeletons were not close relatives but probably linked to the same clan structure. Comparison of their haplotypes with the haplotypes of 8,774 Eurasian individuals suggested a relative specificity and continuity of part of the Yakut mitochondrial gene pool during the last 3 centuries.