Setor Técnico-Científico, Superintendência Regional do Rio Grande do Sul, Polícia Federal, Porto Alegre, Brazil; Escola de Ciências, Pontifícia Universidade Católica do Rio Grande do Sul, Porto Alegre, Brazil; Instituto Nacional de Ciência e Tecnologia INCT Ciências Forenses, Porto Alegre, Brazil.
Escola de Ciências, Pontifícia Universidade Católica do Rio Grande do Sul, Porto Alegre, Brazil.
Forensic Sci Int Genet. 2019 Sep;42:154-164. doi: 10.1016/j.fsigen.2019.07.004. Epub 2019 Jul 9.
The use of Massive Parallel Sequencing (MPS) techniques have been proposed by the forensic community as an alternative to Sanger sequencing methods in routine forensic casework analysis regarding mitochondrial DNA (mtDNA). Interesting features of MPS include high throughput, ability to simultaneously genotype a significant number of samples by barcoding techniques, processing automation, reduced time and costs, among others. Advantages include the capability of generating full mtDNA genome sequences versus usual techniques, usually limited to hypervariable or control regions exclusively. In this work, 96 reference single-source samples from three different Brazilian cities were subjected to full mtDNA genome sequencing by MPS techniques using an early-access version of Precision ID mtDNA Whole Genome Panel on an Ion Torrent PGM platform (Thermo Fisher Scientific, Waltham, MA, USA). Complete, high-quality sequences were obtained and sequencing performance was evaluated via four different metrics. As a subset of evaluated samples have been previously submitted for Sanger sequencing of the control region, a comparative analysis of both methods' results was conducted in order to compare technique adequacy within a forensic context. Even though this study is one of the first to report full mtDNA genome sequences for Brazilian admixed populations, the observed haplotypes exhibit a predominance of Native American and African maternal lineages in the studied sample set, reproducing results described in the literature for control regions only. Interpopulation analysis among Brazilian and 26 worldwide populations was also carried out. The results indicate that MPS-generated full mtDNA genome sequences may have great utility in forensic real casework applications, with a pronounced gain of genetic information and discrimination power provided by coding region evaluation and the enhanced capacity of heteroplasmies determination. Database construction and other relevant factors concerning implementation of such techniques in Brazilian forensic laboratories are also discussed.
大规模平行测序（MPS）技术已被法医界提出，作为常规法医案例分析中替代 Sanger 测序方法的一种选择，用于线粒体 DNA（mtDNA）。MPS 的有趣特点包括高通量、通过条形码技术同时对大量样本进行基因分型的能力、处理自动化、缩短时间和降低成本等。其优点包括能够生成完整的 mtDNA 基因组序列，而通常的技术仅限于高变区或控制区。在这项工作中，96 个来自三个不同巴西城市的参考单源样本通过 MPS 技术进行了全 mtDNA 基因组测序，使用 Ion Torrent PGM 平台上的早期访问版本 Precision ID mtDNA 全基因组面板（Thermo Fisher Scientific，Waltham，MA，USA）。获得了完整的高质量序列，并通过四个不同的指标评估了测序性能。由于评估样本的一部分已经提交用于控制区的 Sanger 测序，因此进行了两种方法结果的比较分析，以便在法医背景下比较技术的充分性。尽管这项研究是首批报告巴西混合人群全 mtDNA 基因组序列的研究之一，但观察到的单倍型在研究样本集中表现出美洲原住民和非洲母系血统的优势，复制了仅在控制区描述的文献结果。还对巴西和 26 个全球人群进行了种群间分析。结果表明，MPS 生成的全 mtDNA 基因组序列可能在法医实际案例工作中有很大的应用价值，通过编码区评估提供了更多的遗传信息和鉴别力，以及增强了异质体确定的能力。还讨论了此类技术在巴西法医实验室实施的数据库构建和其他相关因素。
