International Journal of Legal Medicine

, Volume 127, Issue 2, pp 321–333 | Cite as

Pressure cycling technology (PCT) reduces effects of inhibitors of the PCR

  • Pamela L. Marshall
  • Jonathan L. King
  • Nathan P. Lawrence
  • Alexander Lazarev
  • Vera S. Gross
  • Bruce Budowle
Original Article
First Online:
Received:
Accepted:

Abstract

A common problem in the analysis of forensic human DNA evidence, or for that matter any nucleic acid analysis, is the presence of contaminants or inhibitors. Contaminants may copurify with the DNA, inhibiting downstream PCR or they may present samples effectively as containing fewer templates than exist in the PCR, even when the actual amount of DNA is adequate. Typically, these challenged samples exhibit allele imbalance, allele dropout, and sequence-specific inhibition, leading to interpretational difficulties. Lessening the effects of inhibitors may increase the effective yield of challenged low template copy samples. High pressure may alter some inhibitors and render them less effective at reducing the yield of PCR products. In an attempt to enhance the amplicon yield of inhibited DNA samples, pressure cycling technology was applied to DNA exposed to various concentrations of hematin (0, 1.25, 2.5, 5, and 7 μM) and humic acid (0, 1.25, 2.5, 5, and 7 ng/μL). The effect of high pressure on the inhibitors, and subsequently the PCR process, was assessed by measuring DNA quantity by quantitative PCR and evaluating short tandem repeat typing results. The results support that pressure cycling technology reduces inhibitory effects and thus, in effect, enhances yield of contaminated amplified products of both hematin and humic acid contaminate samples. Based on the results obtained in this study, this method can improve the ability to type challenged or inhibited DNA samples.

Keywords

PCR inhibition Pressure cycling technology Forensic DNA analysis Hematin Humic acid Hi-Flow column 
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Notes

Acknowledgments

This project was supported in part by award no. 2009-DN-BX-K188, awarded by the National Institute of Justice, Office of Justice Programs, US Department of Justice. The opinions, findings, and conclusions or recommendations expressed in this publication/program/exhibition are those of the author(s) and do not necessarily reflect those of the Department of Justice.

Conflict of interest

The authors PLM, JLK, and BB declare that they have no conflict of interest. Authors NPL, AL, and VSG are employed by Pressure Biosciences Incorporated.

Ethical standards

The work described above was performed in accordance with all laws (both Federal and State) that apply to research, researcher conduct, and the protection of human test subjects. We also operate under the guidance of and in accordance with the policies of the UNTHSC Institutional Review Board.

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Copyright information

© Springer-Verlag 2012

Authors and Affiliations

  • Pamela L. Marshall
    • 1
  • Jonathan L. King
    • 1
  • Nathan P. Lawrence
    • 2
  • Alexander Lazarev
    • 2
  • Vera S. Gross
    • 2
  • Bruce Budowle
    • 1
  1. 1.Institute of Applied Genetics, Department of Forensic and Investigative GeneticsUniversity of North Texas Health Science CenterFort WorthUSA
  2. 2.Pressure BioSciences IncorporatedSouth EastonUSA

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