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Sequencing the hypervariable regions of human mitochondrial DNA using massively parallel sequencing: Enhanced data acquisition for DNA samples encountered in forensic testing.
Leg Med (Tokyo). 2015 Mar; 17(2):123-7.LM

Abstract

Mitochondrial DNA testing is a useful tool in the analysis of forensic biological evidence. In cases where nuclear DNA is damaged or limited in quantity, the higher copy number of mitochondrial genomes available in a sample can provide information about the source of a sample. Currently, Sanger-type sequencing (STS) is the primary method to develop mitochondrial DNA profiles. This method is laborious and time consuming. Massively parallel sequencing (MPS) can increase the amount of information obtained from mitochondrial DNA samples while improving turnaround time by decreasing the numbers of manipulations and more so by exploiting high throughput analyses to obtain interpretable results. In this study 18 buccal swabs, three different tissue samples from five individuals, and four bones samples from casework were sequenced at hypervariable regions I and II using STS and MPS. Sample enrichment for STS and MPS was PCR-based. Library preparation for MPS was performed using Nextera® XT DNA Sample Preparation Kit and sequencing was performed on the MiSeq™ (Illumina, Inc.). MPS yielded full concordance of base calls with STS results, and the newer methodology was able to resolve length heteroplasmy in homopolymeric regions. This study demonstrates short amplicon MPS of mitochondrial DNA is feasible, can provide information not possible with STS, and lays the groundwork for development of a whole genome sequencing strategy for degraded samples.

Authors+Show Affiliations

Department of Molecular and Medical Genetics, Institute of Applied Genetics, University of North Texas Health Science Center, 3500 Camp Bowie Blvd, Fort Worth, TX 76107, USA.Department of Molecular and Medical Genetics, Institute of Applied Genetics, University of North Texas Health Science Center, 3500 Camp Bowie Blvd, Fort Worth, TX 76107, USA.Department of Molecular and Medical Genetics, Institute of Applied Genetics, University of North Texas Health Science Center, 3500 Camp Bowie Blvd, Fort Worth, TX 76107, USA.Department of Molecular and Medical Genetics, Institute of Applied Genetics, University of North Texas Health Science Center, 3500 Camp Bowie Blvd, Fort Worth, TX 76107, USA.Department of Molecular and Medical Genetics, Institute of Applied Genetics, University of North Texas Health Science Center, 3500 Camp Bowie Blvd, Fort Worth, TX 76107, USA; Center of Excellence in Genomic Medicine Research (CEGMR), King Abdulaziz University, Jeddah, Saudi Arabia. Electronic address: bruce.budowle@unthsc.edu.

Pub Type(s)

Journal Article

Language

eng

PubMed ID

25459369

Citation

Davis, Carey, et al. "Sequencing the Hypervariable Regions of Human Mitochondrial DNA Using Massively Parallel Sequencing: Enhanced Data Acquisition for DNA Samples Encountered in Forensic Testing." Legal Medicine (Tokyo, Japan), vol. 17, no. 2, 2015, pp. 123-7.
Davis C, Peters D, Warshauer D, et al. Sequencing the hypervariable regions of human mitochondrial DNA using massively parallel sequencing: Enhanced data acquisition for DNA samples encountered in forensic testing. Leg Med (Tokyo). 2015;17(2):123-7.
Davis, C., Peters, D., Warshauer, D., King, J., & Budowle, B. (2015). Sequencing the hypervariable regions of human mitochondrial DNA using massively parallel sequencing: Enhanced data acquisition for DNA samples encountered in forensic testing. Legal Medicine (Tokyo, Japan), 17(2), 123-7. https://doi.org/10.1016/j.legalmed.2014.10.004
Davis C, et al. Sequencing the Hypervariable Regions of Human Mitochondrial DNA Using Massively Parallel Sequencing: Enhanced Data Acquisition for DNA Samples Encountered in Forensic Testing. Leg Med (Tokyo). 2015;17(2):123-7. PubMed PMID: 25459369.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Sequencing the hypervariable regions of human mitochondrial DNA using massively parallel sequencing: Enhanced data acquisition for DNA samples encountered in forensic testing. AU - Davis,Carey, AU - Peters,Dixie, AU - Warshauer,David, AU - King,Jonathan, AU - Budowle,Bruce, Y1 - 2014/10/25/ PY - 2014/06/22/received PY - 2014/10/03/revised PY - 2014/10/05/accepted PY - 2014/12/3/entrez PY - 2014/12/3/pubmed PY - 2015/11/18/medline KW - HVI KW - HVII KW - Heteroplasmy KW - Massively parallel sequencing KW - Mitochondrial DNA KW - Sanger type sequencing SP - 123 EP - 7 JF - Legal medicine (Tokyo, Japan) JO - Leg Med (Tokyo) VL - 17 IS - 2 N2 - Mitochondrial DNA testing is a useful tool in the analysis of forensic biological evidence. In cases where nuclear DNA is damaged or limited in quantity, the higher copy number of mitochondrial genomes available in a sample can provide information about the source of a sample. Currently, Sanger-type sequencing (STS) is the primary method to develop mitochondrial DNA profiles. This method is laborious and time consuming. Massively parallel sequencing (MPS) can increase the amount of information obtained from mitochondrial DNA samples while improving turnaround time by decreasing the numbers of manipulations and more so by exploiting high throughput analyses to obtain interpretable results. In this study 18 buccal swabs, three different tissue samples from five individuals, and four bones samples from casework were sequenced at hypervariable regions I and II using STS and MPS. Sample enrichment for STS and MPS was PCR-based. Library preparation for MPS was performed using Nextera® XT DNA Sample Preparation Kit and sequencing was performed on the MiSeq™ (Illumina, Inc.). MPS yielded full concordance of base calls with STS results, and the newer methodology was able to resolve length heteroplasmy in homopolymeric regions. This study demonstrates short amplicon MPS of mitochondrial DNA is feasible, can provide information not possible with STS, and lays the groundwork for development of a whole genome sequencing strategy for degraded samples. SN - 1873-4162 UR - https://www.unboundmedicine.com/medline/citation/25459369/Sequencing_the_hypervariable_regions_of_human_mitochondrial_DNA_using_massively_parallel_sequencing:_Enhanced_data_acquisition_for_DNA_samples_encountered_in_forensic_testing_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S1344-6223(14)00158-8 DB - PRIME DP - Unbound Medicine ER -