title: Full mtGenome reference data: development and characterization of 588 forensic-quality haplotypes representing three U.S. populations.
abstract: Though investigations into the use of massively parallel sequencing technologies for the generation of complete mitochondrial genome (mtGenome) profiles from difficult forensic specimens are well underway in multiple laboratories, the high quality population reference data necessary to support full mtGenome typing in the forensic context are lacking. To address this deficiency, we have developed 588 complete mtGenome haplotypes, spanning three U.S. population groups (African American, Caucasian and Hispanic) from anonymized, randomly-sampled specimens. Data production utilized an 8-amplicon, 135 sequencing reaction Sanger-based protocol, performed in semi-automated fashion on robotic instrumentation. Data review followed an intensive multi-step strategy that included a minimum of three independent reviews of the raw data at two laboratories; repeat screenings of all insertions, deletions, heteroplasmies, transversions and any additional private mutations; and a check for phylogenetic feasibility. For all three populations, nearly complete resolution of the haplotypes was achieved with full mtGenome sequences: 90.3-98.8% of haplotypes were unique per population, an improvement of 7.7-29.2% over control region sequencing alone, and zero haplotypes overlapped between populations. Inferred maternal biogeographic ancestry frequencies for each population and heteroplasmy rates in the control region were generally consistent with published datasets. In the coding region, nearly 90% of individuals exhibited length heteroplasmy in the 12418-12425 adenine homopolymer; and despite a relatively high rate of point heteroplasmy (23.8% of individuals across the entire molecule), coding region point heteroplasmies shared by more than one individual were notably absent, and transversion-type heteroplasmies were extremely rare. The ratio of nonsynonymous to synonymous changes among point heteroplasmies in the protein-coding genes (1:1.3) and average pathogenicity scores in comparison to data reported for complete substitutions in previous studies seem to provide some additional support for the role of purifying selection in the evolution of the human mtGenome. Overall, these thoroughly vetted full mtGenome population reference data can serve as a standard against which the quality and features of future mtGenome datasets (especially those developed via massively parallel sequencing) may be evaluated, and will provide a solid foundation for the generation of complete mtGenome haplotype frequency estimates for forensic applications.
authors.affiliation: Armed Forces DNA Identification Laboratory, 115 Purple Heart Dr., Dover AFB, DE 19902, United States; American Registry of Pathology, 15245 Shady Grove Rd, Suite 335, Rockville, MD 20850, United States; University of Maryland, College Park, 8082 Baltimore Ave., College Park, MD 20740, United States. Electronic address: rebecca.s.just.ctr@mail.mil.
authors.forename: Rebecca S
authors.initials: RS
authors.lastname: Just
authors.affiliation: Armed Forces DNA Identification Laboratory, 115 Purple Heart Dr., Dover AFB, DE 19902, United States; American Registry of Pathology, 15245 Shady Grove Rd, Suite 335, Rockville, MD 20850, United States.
authors.forename: Melissa K
authors.initials: MK
authors.lastname: Scheible
authors.affiliation: Armed Forces DNA Identification Laboratory, 115 Purple Heart Dr., Dover AFB, DE 19902, United States; American Registry of Pathology, 15245 Shady Grove Rd, Suite 335, Rockville, MD 20850, United States.
authors.forename: Spence A
authors.initials: SA
authors.lastname: Fast
authors.affiliation: Armed Forces DNA Identification Laboratory, 115 Purple Heart Dr., Dover AFB, DE 19902, United States; American Registry of Pathology, 15245 Shady Grove Rd, Suite 335, Rockville, MD 20850, United States.
authors.forename: Kimberly
authors.initials: K
authors.lastname: Sturk-Andreaggi
authors.affiliation: Institute of Legal Medicine, Innsbruck Medical University, Müllerstrasse 44, Innsbruck, Austria.
authors.forename: Alexander W
authors.initials: AW
authors.lastname: Röck
authors.affiliation: Armed Forces DNA Identification Laboratory, 115 Purple Heart Dr., Dover AFB, DE 19902, United States; American Registry of Pathology, 15245 Shady Grove Rd, Suite 335, Rockville, MD 20850, United States.
authors.forename: Jocelyn M
authors.initials: JM
authors.lastname: Bush
authors.affiliation: Armed Forces DNA Identification Laboratory, 115 Purple Heart Dr., Dover AFB, DE 19902, United States; American Registry of Pathology, 15245 Shady Grove Rd, Suite 335, Rockville, MD 20850, United States.
authors.forename: Jennifer L
authors.initials: JL
authors.lastname: Higginbotham
authors.affiliation: Armed Forces DNA Identification Laboratory, 115 Purple Heart Dr., Dover AFB, DE 19902, United States; American Registry of Pathology, 15245 Shady Grove Rd, Suite 335, Rockville, MD 20850, United States.
authors.forename: Michelle A
authors.initials: MA
authors.lastname: Peck
authors.affiliation: Armed Forces DNA Identification Laboratory, 115 Purple Heart Dr., Dover AFB, DE 19902, United States; American Registry of Pathology, 15245 Shady Grove Rd, Suite 335, Rockville, MD 20850, United States.
authors.forename: Joseph D
authors.initials: JD
authors.lastname: Ring
authors.affiliation: Institute of Legal Medicine, Innsbruck Medical University, Müllerstrasse 44, Innsbruck, Austria.
authors.forename: Gabriela E
authors.initials: GE
authors.lastname: Huber
authors.affiliation: Institute of Legal Medicine, Innsbruck Medical University, Müllerstrasse 44, Innsbruck, Austria.
authors.forename: Catarina
authors.initials: C
authors.lastname: Xavier
authors.affiliation: Institute of Legal Medicine, Innsbruck Medical University, Müllerstrasse 44, Innsbruck, Austria.
authors.forename: Christina
authors.initials: C
authors.lastname: Strobl
authors.affiliation: Armed Forces DNA Identification Laboratory, 115 Purple Heart Dr., Dover AFB, DE 19902, United States; American Registry of Pathology, 15245 Shady Grove Rd, Suite 335, Rockville, MD 20850, United States.
authors.forename: Elizabeth A
authors.initials: EA
authors.lastname: Lyons
authors.affiliation: Armed Forces DNA Identification Laboratory, 115 Purple Heart Dr., Dover AFB, DE 19902, United States; American Registry of Pathology, 15245 Shady Grove Rd, Suite 335, Rockville, MD 20850, United States.
authors.forename: Toni M
authors.initials: TM
authors.lastname: Diegoli
authors.affiliation: Institute of Legal Medicine, Innsbruck Medical University, Müllerstrasse 44, Innsbruck, Austria.
authors.forename: Martin
authors.initials: M
authors.lastname: Bodner
authors.affiliation: Institute of Legal Medicine, Innsbruck Medical University, Müllerstrasse 44, Innsbruck, Austria.
authors.forename: Liane
authors.initials: L
authors.lastname: Fendt
authors.affiliation: Institute of Legal Medicine, Innsbruck Medical University, Müllerstrasse 44, Innsbruck, Austria.
authors.forename: Petra
authors.initials: P
authors.lastname: Kralj
authors.affiliation: Institute of Legal Medicine, Innsbruck Medical University, Müllerstrasse 44, Innsbruck, Austria.
authors.forename: Simone
authors.initials: S
authors.lastname: Nagl
authors.affiliation: Institute of Legal Medicine, Innsbruck Medical University, Müllerstrasse 44, Innsbruck, Austria.
authors.forename: Daniela
authors.initials: D
authors.lastname: Niederwieser
authors.affiliation: Institute of Legal Medicine, Innsbruck Medical University, Müllerstrasse 44, Innsbruck, Austria.
authors.forename: Bettina
authors.initials: B
authors.lastname: Zimmermann
authors.affiliation: Institute of Legal Medicine, Innsbruck Medical University, Müllerstrasse 44, Innsbruck, Austria; Penn State Eberly College of Science, 517 Thomas Building, University Park, PA 16802, United States.
authors.forename: Walther
authors.initials: W
authors.lastname: Parson
authors.affiliation: Armed Forces DNA Identification Laboratory, 115 Purple Heart Dr., Dover AFB, DE 19902, United States; American Registry of Pathology, 15245 Shady Grove Rd, Suite 335, Rockville, MD 20850, United States.
authors.forename: Jodi A
authors.initials: JA
authors.lastname: Irwin
mesh_terms: D053803:Forensic Genetics; D054629:Genome, Mitochondrial; D006239:Haplotypes; D006801:Humans; D014481:United States
keywords: Haplogroup; Heteroplasmy; Reference population database; Sequencing; mtDNA; mtGenome