Research Article

Resolving a human identification case for the Rio de Janeiro Police with massively parallel sequencing of mtDNA using a proposed pipeline

Published: March 31, 2021
Genet. Mol. Res. 20(1): GMR18757 DOI:


The forensic community is concerned about the quality control of mitotypes reported, as probability estimates may be biased to the disadvantage of a suspect when there is incomplete data. This is particularly important when using new technology such as Massively Parallel Sequencing (MPS), which performs simultaneous analysis of millions of DNA fragments, generating vast amounts of data from small amounts of sample, providing more extensive genetic information. The use of MPS for complex forensic evidence analysis could save time and resources, and provide reliable data for extremely degraded DNA samples. However, data analysis methods can be challenging to implement in a forensic labs routine. Here, we propose a reliable and straightforward pipeline for mtDNA data analysis in forensic samples using MPS, and we used this pipeline to solve a human identification case by Rio de Janeiro Police who had recovered a severely putrefied body that could not be identified by conventional autosomal STR analysis. Mitochondrial DNA (mtDNA) from samples were sequenced on Ion Torrent PGM. Raw data from the sequencing was analyzed in two main pipeline protocols: quality control and forensic analysis. The first includes the analyses of raw data and contamination control. The second generates haplotypes, allowing haplogroup classification, and biogeographic inference, as well as verification of a matrilineal relationship. This straightforward freeware pipeline for mtDNA data analysis should facilitate the implementation of this type of system in forensic lab routine. Using our pipeline, the definition of the samples being of African origin was more precise when considering the whole mtGenome and not only the control region, which is usually used for routine forensic analysis. The final analysis was consistent with the existence of a matrilineal relationship between the alleged son (bone sample recovered from the sea) and the mother.

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