20.500.12556/RUL-154409 Comparison of the optimal and suboptimal quantity of mitotype libraries using next-generation sequencing Optimizing analysis parameters and sample input is crucial in forensic genetics methods to generate reliable results, and even more so when working with muti-copy mitochondrial DNA (mtDNA) and low-quality samples. This study compared mitotypes based on next-generation sequencing (NGS) results derived from the same samples at two different sequencing library concentrations—30 pM and 0.3 pM. Thirty femur samples from the Second World War were used as a model for poorly preserved DNA. Quantitative PCR (qPCR) method targeting 113 bp long fragment was employed to assess the quantity of mitogenomes. HID Ion Chef™ Instrument with Precision ID mtDNA Control Region Panel was used for library preparation and templating. Sequencing was performed with Ion GeneStudio™ S5 System. Reference haplotypes were determined from sequencing samples at 30 pM library input. Haplotypes were compared between optimal (30 pM) and suboptimal (0.3 pM) library inputs. Often the difference in haplotypes was length heteroplasmy, which in line with other studies shows that this type of variant is not reliable for interpretation in forensics. Excluding length variants at positions 573, 309, and 16,193, 56.7% of the samples matched, and in two samples, no sequence was obtained at suboptimal library input. The rest of the samples differed between optimal and suboptimal library input. To conclude, genotyping and analyzing low-quantity libraries derived from low-quality aged skeletonized human remains therefore must be done with caution in forensic genetics casework. haplotype library input mitogenome next-generation sequencing Second World War haplotip vnos v knjižnico mitogenom sekvenciranje naslednje generacije druga svetovna vojna true false true Angleški jezik Slovenski jezik Članek v reviji 2024-02-13 14:20:52 2024-02-13 14:20:56 2024-02-14 03:46:39 0000-00-00 00:00:00 2024 0 0 Str. 395–400 iss. 2 Vol. 138 2024 0000-00-00 Zaloznikova Objavljeno NiDoloceno 0000-00-00 0000-00-00 0000-00-00 616:340.6 1437-1596 10.1007/s00414-023-03099-7 167475971 s00414-023-03099-7.pdf s00414-023-03099-7.pdf 1 B1725E597F78F20AA6E82C0F49C1A300 d35998c9a8de4fa556da669e097b5c47117278a5573d856cc8c1de80ab0bcec5 f225d9ba-ca72-11ee-a430-0050569b8976 https://repozitorij.uni-lj.si/Dokument.php?lang=slv&id=180824 https://link.springer.com/article/10.1007/s00414-023-03099-7 1 b91878b7-ca72-11ee-a430-0050569b8976 https://repozitorij.uni-lj.si/Dokument.php?lang=slv&id=180823 Medicinska fakulteta 0 0 0