In an attempt to improve the amplicon yield of inhibited DNA samples, the current project applied pressure cycling technology 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).
A common problem in the analysis of forensic human DNA evidence or 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 the current project, 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 indicate that pressure cycling technology reduces inhibitory effects and thus, in effect, improves 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. (publisher abstract modified)
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