Maximizing DNA Profiling Success From Sub-Optimal Quantities of DNA: A Staged Approach

• NCJ Number: 226999
• Journal: Forensic Science International: Genetics Volume: 3 Issue: 2 Dated: March 2009 Pages: 128-137
• Author(s): Amy D. Roeder; Paul Elsmore; Matt Greenhalgh; Andrew McDonald
• Date Published: March 2009
• Length: 10 pages
• Publication Format: Article
• Publication Type: Report (Technical); Report (Study/Research)
• Language: English
• Country: Ireland

Annotation

This study evaluated the effects of additional thermocycle numbers and electro-injection of increased quantities of PCR product (termed Enhancement) on both SGM Plus and Identifiler profiles generated from suboptimal quantities of DNA.

Abstract

The study found that the increased thermocycle number and PCR product Enhancement techniques described in this paper allowed for adaptable processing of each sample. In this staged approach, the sample is assessed after DNA quantification in order to determine whether PCR should be performed using 28 or 30 thermocycles. The genetic profile is then assessed in order to determine whether to conduct Phase One Enhancement and which thermocycle number to use for the replicate PCR. Profiles generated following Phase One Enhancement can then be used to determine whether to proceed to Phase Two Enhancement. This staged approach allows for an optimal profile to be generated from each sample, while conserving DNA extract and minimizing costs. In order to evaluate the increased thermocycle number and Enhancement methods for generating accurate genetic profiles from samples that contained suboptimal amounts of DNA, four sets of experiments were conducted. All reactions were conducted with either AmpFlSTR SGM Plus or AmpFlSTR Identifiler PCR Amplification Kits. PCRs were prepared and amplified according to the manufacturer’s instructions, with the exceptions that the SGM Plus reactions were set up in 25 ml volumes, and the amounts of DNA added were generally lower than those recommended by the manufacturer. Also, the number of thermocycles was increased to 30 or 34 for some reactions. PCR products from putative single-source samples that gave partial or no profiles were enhanced, i.e., subjected to the Enhancement process. The Phase One and Phase Two Enhancement procedures are described, along with the samples, DNA extraction and quantification, amplification and electrophoresis of PCR products, and profile analysis. 10 tables, 3 figures, and 11 references