Direct Comparison of Post-28-Cycle PCR Purification and Modified Capillary Electrophoresis Methods With the 34-Cycle "Low Copy Number" (LCN) Method for Analysis of Trace Forensic DNA Samples

• NCJ Number: 225049
• Journal: Forensic Science International: Genetics Volume: 2 Issue: 4 Dated: September 2008 Pages: 318-328
• Author(s): Luke Forster; Jim Thomson; Stefan Kutranov
• Date Published: September 2008
• Length: 11 pages
• Publication Format: Article
• Publication Type: Report (Study/Research)
• Language: English
• Country: Ireland

Annotation

This article reviewed the characteristics of the DNA profiles obtained by optimizing post-PCR purification techniques applied after only 28 cycles of PCR, as well as the use of modified capillary electrophoresis injection conditions; these profiles were compared with those obtained on the same samples after 34-cycle PCR.

Abstract

The 28 cycles of PCR displayed comparable sensitivity to 34-cycle PCR in terms of the number of profiles with evidence of DNA and the number of allelic peaks per profile. Improved peak height and area magnitude with some sample types were observed with the 28-cycle PCR. Certain parameters reported to be adversely affected in 34-cycle “low copy number” (LCN) investigations--such as non-donor allele peaks and increased stutter peak ratio--were reduced by the 28-cycle PCR. There are a number of advantages for trace samples in progressing from the 28-cycle process to the post-PCR enhancement methods in order to increase signal from trace DNA samples compared to the 34-cycle PCR method. These include reduced sample consumption, reduced number of PCR amplifications required, and a staged approach to sample processing and profile interpretation. The study used the Qiagen MinElute column in order to both clean-up and concentrate the AmpF/STR SGM Plus PCR product, which was amplified at the manufacturer’s recommended volume of 50 ml, which is at least twice the volume used in other LCN studies, allowing for a subsequently greater concentration of product. An increased volume of the product was then added to the Hi-Di Formamide mix, increasing the amount of product injected into the capillary. Lastly, the injection time and voltage were increased. These factors together greatly increased the amount of amplified DNA injected into the capillary compared to untreated post-PCR reaction mix. 9 tables, 3 figures, and 18 references