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Production and certification of NIST Standard Reference Material 2372 Human DNA Quantitation Standard

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Abstract

Modern highly multiplexed short tandem repeat (STR) assays used by the forensic human-identity community require tight control of the initial amount of sample DNA amplified in the polymerase chain reaction (PCR) process. This, in turn, requires the ability to reproducibly measure the concentration of human DNA, [DNA], in a sample extract. Quantitative PCR (qPCR) techniques can determine the number of intact stretches of DNA of specified nucleotide sequence in an extremely small sample; however, these assays must be calibrated with DNA extracts of well-characterized and stable composition. By 2004, studies coordinated by or reported to the National Institute of Standards and Technology (NIST) indicated that a well-characterized, stable human DNA quantitation certified reference material (CRM) could help the forensic community reduce within- and among-laboratory quantitation variability. To ensure that the stability of such a quantitation standard can be monitored and that, if and when required, equivalent replacement materials can be prepared, a measurement of some stable quantity directly related to [DNA] is required. Using a long-established conventional relationship linking optical density (properly designated as decadic attenuance) at 260 nm with [DNA] in aqueous solution, NIST Standard Reference Material (SRM) 2372 Human DNA Quantitation Standard was issued in October 2007. This SRM consists of three quite different DNA extracts: a single-source male, a multiple-source female, and a mixture of male and female sources. All three SRM components have very similar optical densities, and thus very similar conventional [DNA]. The materials perform very similarly in several widely used gender-neutral assays, demonstrating that the combination of appropriate preparation methods and metrologically sound spectrophotometric measurements enables the preparation and certification of quantitation [DNA] standards that are both maintainable and of practical utility.

NIST Standard Reference Material (SRM) 2372 Human Quantitation Standard

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Acknowledgements

We thank all of the forensic scientists, supervisors, and forensic DNA testing facilities who contributed their expertise, time, and material resources to the interlaboratory assessment. SRM 2372 would not exist except for their encouragement, advice, and willing cooperation with our sometimes seemingly outré requests. The development of SRM 2372 was funded in part by the National Institute of Justice through an interagency agreement with the NIST Office of Law Enforcement Standards.

Disclaimer

Certain commercial equipment, instruments, and materials are identified in order to specify experimental procedures as completely as possible. In no case does such identification imply a recommendation or endorsement by the National Institute of Standards and Technology nor does it imply that any of the materials, instruments, or equipment identified are necessarily the best available for the purpose.

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Authors and Affiliations

  1. Biochemical Science Division, National Institute of Standards and Technology, 100 Bureau Drive, MS 8311, Gaithersburg, MD, 20899-8311, USA

    Margaret C. Kline, John C. Travis, Melody V. Smith, Janette W. Redman, Peter M. Vallone, Amy E. Decker & John M. Butler

  2. Analytical Chemistry Division, National Institute of Standards and Technology, 100 Bureau Drive, MS 8390, Gaithersburg, MD, 20899-8390, USA

    David L. Duewer

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  1. Margaret C. Kline
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  2. David L. Duewer
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  6. Peter M. Vallone
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  7. Amy E. Decker
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  8. John M. Butler
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Correspondence to Margaret C. Kline.

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Kline, M.C., Duewer, D.L., Travis, J.C. et al. Production and certification of NIST Standard Reference Material 2372 Human DNA Quantitation Standard. Anal Bioanal Chem 394, 1183–1192 (2009). https://doi.org/10.1007/s00216-009-2782-0

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  • DOI: https://doi.org/10.1007/s00216-009-2782-0

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