Volume 46, Issue 2 (March 2001)

    Improved MtDNA Sequence Analysis of Forensic Remains Using a “Mini-Primer Set” Amplification Strategy

    (Received 14 February 2000; accepted 15 May 2000)

    CODEN: JFSOAD

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    Abstract

    Mitochondrial DNA (mtDNA) analysis of highly degraded skeletal remains is often used for forensic identification due largely to the high genome copy number per cell. Literature from the “ancient DNA” field has shown that highly degraded samples contain populations of intact DNA molecules that are severely restricted in size (1–4). Hand et al. have demonstrated the targeting and preferential amplification of authentic human DNA sequences with small amplicon products of 150 bp or less (1,2). Given this understanding of ancient DNA preservation and amplification, we report an improved approach to forensic mtDNA analysis of hypervariable regions 1 and 2 (HV1/HV2) in highly degraded specimens. This “mini-primer set” (MPS) amplification strategy consists of four overlapping products that span each of the HV regions and range from 126 to 170 bp, with an average size of 141 bp. For this study, 11 extracts representing a range of sample quality were prepared from nonprobative forensic specimens. We demonstrate a significant increase in MPS amplification success when compared to testing methods using ∼250 bp amplicons. Further, 16 of 17 independent amplifications previously “unreported” due to mixed sequences provided potentially reportable sequence data from a single, authentic template with MPS testing.


    Author Information:

    Huffine, EF
    DNA Program Director, Roche Molecular Systems, Alameda, CA

    Ryan, JH
    DNA Research Technologist II, Technical Leader, MtDNA Section, Scientific Laboratory Director, and Chief Scientiest, The Armed Forces DNA Identification Laboratory, Rockville,

    Holland, MM
    DNA Research Technologist II, Technical Leader, MtDNA Section, Scientific Laboratory Director, and Chief Scientiest, The Armed Forces DNA Identification Laboratory, Rockville,

    Gabriel, MN
    DNA Research Technologist II, Technical Leader, MtDNA Section, Scientific Laboratory Director, and Chief Scientiest, The Armed Forces DNA Identification Laboratory, Rockville,

    Parsons, TJ
    DNA Research Technologist II, Technical Leader, MtDNA Section, Scientific Laboratory Director, and Chief Scientiest, The Armed Forces DNA Identification Laboratory, Rockville,


    Stock #: JFS14957J

    ISSN: 0022-1198

    DOI: 10.1520/JFS14957J

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    Title Improved MtDNA Sequence Analysis of Forensic Remains Using a “Mini-Primer Set” Amplification Strategy
    Symposium , 0000-00-00
    Committee E30