Volume 2, Issue 1 , Pages 322-323, December 2009
MtDNA SNP analysis in a Central Portuguese population
Article Outline
- Abstract
- 1. Introduction
- 2. Material and methods
- 3. Results and discussion
- 4. Conclusion
- Conflict of interest
- References
- Copyright
Abstract
Mitochondrial DNA (mtDNA) has a great potential in forensic and population genetics, allowing the identification of population origin of individuals and related trace samples. In order to apply this methodology as inclusion evidence in forensic practice it is necessary to determine the population genetic structure. Here, we present haplogroup discrimination of a Central Portuguese population using 16 mtSNPs. Haplogroups in the Central Portuguese population were determined with the following distribution: Hg H: 40.19%, Hg V: 4.90%; Hg HV: 4.90%, Hg U: 13.72%, Hg K: 5.88%; Hg T: 10.78%; Hg J: 9.80%, Hg I: 1.96%, Hg X: 2.94% and others: 4.90%, being this distribution in accordance with other European populations.
Keywords: Mitochondrial DNA, Coding region SNPs, SNaPshot, West Eurasian, Haplogroup
1. Introduction
Mitochondrial DNA (mtDNA) has been used extensively in the study of the origin and dispersal of modern humans, primarily not only due to its matrilineal inheritance, not suffering recombination, but also due to its mutation rate that leaded to the sequential mutation accumulation through maternal lineages that diverged as human populations colonized different parts of the globe [1].
Western Eurasians may be classified into 9 major haplogroups and several sub-haplogroups: H, I, J, K, T, U, V, W, and X [1], [2]. These haplogroups do not show an uniform distribution, with Hg H being the most frequent, reaching up to 60% in European populations [3]. The information obtained with mtDNA control region polymorphisms (HVS-1 and HVS-2) [4], [5] is insufficient to discriminate between samples belonging to the same haplogroup, so several protocols to analyse mtDNA SNPs from coding region were developed in order to achieve more information [6], [7], [8], [9], since they are distributed over the entire mitochondrial genome. Here, we present haplogroup discrimination of a Central Portuguese population using 16 mtSNPs.
2. Material and methods
DNA was extracted from 102 bloodstains using Chelex 100 method [10] and DNA quantification was achieved with Quantifiler Human DNA Quantification Kit at an ABI Prism7000 Sequence Detection System (AB).
Sixteen mtSNP were amplified in two multiplex reactions as described previously [8], [11]. Multiplex I comprised the SNP target sites G709A, G1719A, G3010A, C7028T, A11251G, G12372A, T14798C and C15904T. Multiplex II comprised the SNP target sites A1811G, T6365C, T6776C, G8251A, G8697A, G9055A, G13708A and C14766T. Previous to minisequencing reaction, PCR product was treated with Exo-SAP (USB) in order to remove unincorporated dNTPs and excess primers using an extended protocol. Minisequencing reaction was achieved using SNaPshot kit (AB) [8]. Unincorporated ddNTPs were removed with a SAP treatment (Roche). Electrophoresis was undertaken on an ABI PRISM 3130 Genetic Analyzer using Liz-120 (AB) as internal size standard. Analysis was done using GeneMapper ID v.3.2 software. Samples were typed and assigned into the nine major West European Caucasian haplogroups according to known phylogenetic network of the mitochondrial genome.
3. Results and discussion
All 102 samples were successfully typed with the two multiplexes. The haplotypes found in this study are summarized in Table 1. From the 9 major West European Caucasian haplogroups only 8 were observed in our sample, since none of the analysed individuals belonged to Hg W. The most common haplotype found was a profile assigned to haplogroup H1 (20.6%), presenting the 3010A mutation. Overall, 20 haplotype motifs were found with 8 being unique. Five individuals did not fit into predefined haplogroups and were classified as “undetermined”. Haplogroup H accounted for 40.2% of the population, being 10.8% from H3 and 21.6% from H1. The remaining individuals were classified as H*.
Table 1. MtDNA coding region haplotype motifs in 102 individuals from Central Portugal.
| Haplogroup | Haplotype motif | N | Freq (%) |
|---|---|---|---|
| HV* | 7028T | 4 | 3.92 |
| HV* | 7028T–13708A | 1 | 0.98 |
| V* | 7028T–15904T | 5 | 4.90 |
| H* | None | 8 | 7.84 |
| H3 | 6776C | 11 | 10.78 |
| H1 | 3010A | 21 | 20.59 |
| H1 | 3010A–8251A | 1 | 0.98 |
| U* | 7028T–12372A–14766T | 10 | 9.80 |
| U2′3′4′7′8 | 1811G–7028T–12372A–14766T | 4 | 3.92 |
| K* | 1811G–7028T–9055A–12372A–14766T–14789C | 6 | 5.88 |
| T* | 709A–7028T–8697A–11251G–14766T | 11 | 10.78 |
| J* | 7028T–11251G–13708A–14766T | 1 | 0.98 |
| J1 | 3010A–7028T–11251G–13708A–14766T | 1 | 0.98 |
| J1c | 3010A–7028T–11251G–13708A–14766T–14789C | 8 | 7.84 |
| I*/N1e | 1719A–7028T–8251A–14766T | 1 | 0.98 |
| I*/N1e | 1719A–7028T–8251A–14766T–15904G | 1 | 0.98 |
| X2b’d | 1719A–13708A–14766T | 2 | 1.96 |
| X2 | 1719A––14766T | 1 | 0.98 |
| Undet. | 7028T–14766T | 4 | 3.92 |
| Undet. | 3010A–7028T–14766T | 1 | 0.98 |
Haplogroups from the Central Portuguese population were determined with the following distribution: Hg H: 40.19%, Hg V: 4.90%; Hg HV: 4.90%, Hg U: 13.72%, Hg K: 5.88%; Hg T: 10.78%; Hg J: 9.80%, Hg I: 1.96%, Hg X: 2.94% and undetermined: 4.90%, being this distribution in accordance with other European populations.
4. Conclusion
MtDNA SNPs in spite of being lineage markers not capable of genetic individualization, are extremely useful in forensic genetics and can be used not only as a screening method for large volume cases but also as a technique to differentiate among samples with the same HVS-1 and HVS-2 haplotype.
Conflict of interest
None.
References
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PII: S1875-1768(09)00109-7
doi:10.1016/j.fsigss.2009.08.093
© 2009 Elsevier Ireland Ltd. All rights reserved.
Volume 2, Issue 1 , Pages 322-323, December 2009
