An Investigation of PCR Inhibition Using Plexor®‐Based Quantitative PCR and Short Tandem Repeat Amplification123. Issue 6 (3rd September 2014)

• Record Type: Journal Article
• Authors: Thompson, Robyn E.; Duncan, George; McCord, Bruce R.
• Is Part Of: Journal of forensic sciences. Volume 59:Issue 6(2014)
• Page Start: 1517
• Page End: 1529
• Publication Date: 2014-09-03
• Subjects: Medical jurisprudence -- Periodicals; Forensic sciences -- Periodicals; Forensic Medicine -- Periodicals; Gerechtelijke geneeskunde; Gerechtelijke chemie; Gerechtelijke psychiatrie; 363.2505
• DOI: 10.1111/1556-4029.12556
• Languages: English
• ISSNs: 0022-1198

Abstract

<abstract abstract-type="main" id="jfo12556-abs-0001"> <title>Abstract</title> <p>A common problem in forensic DNA typing is PCR inhibition resulting in allele dropout and peak imbalance. In this paper, we have utilized the Plexor^® real‐time PCR quantification kit to evaluate PCR inhibition. This is performed by adding increasing concentrations of various inhibitors and evaluating changes in melt curves and PCR amplification efficiencies. Inhibitors examined included calcium, humic acid, collagen, phenol, tannic acid, hematin, melanin, urea, bile salts, EDTA, and guanidinium thiocyanate. Results were plotted and modeled using mathematical simulations. In general, we found that PCR inhibitors that bind DNA affect melt curves and C_T takeoff points while those that affect the <italic>Taq</italic> polymerase tend to affect the slope of the amplification curve. Mixed mode effects were also visible. Quantitative PCR results were then compared with subsequent STR amplification using the PowerPlex^® 16 HS System. The overall results demonstrate that real‐time PCR can be an effective method to evaluate PCR inhibition and predict its effects on subsequent STR amplifications.</p> </abstract>