Enhancing Trace DNA Profile Recovery in Forensic Casework Using the Amplicon RX Post-PCR Clean-up Kit

Abstract

This study evaluated the effectiveness of the amplicon RX post-PCR clean-up kit in enhancing trace DNA profile recovery from forensic casework samples amplified using the GlobalFiler PCR amplification kit. The impact of post-PCR clean-up on allele recovery and signal intensity was assessed in both trace casework samples and control samples across a range of DNA concentrations. The results showed that the amplicon RX method significantly improved allele recovery compared to the 29-cycle protocol (p = 8.30 × 10−12) and achieved slightly better results than the 30-cycle protocol (p = 0.019). Additionally, the Amplicon RX method demonstrated a significant increase in signal intensity (p = 2.70 × 10−4), reflecting improved sensitivity in detecting trace DNA profiles compared to the 30-cycle protocol. In the evaluation of control samples, the amplicon RX method consistently outperformed both the 29- and 30-cycle protocols, especially at lower DNA concentrations (D3: 0.001 ng/µL). While the performance of all methods declined at the lowest concentration (D4: 0.0001 ng/µL), the Amplicon RX method still demonstrated superior allele recovery (p = 0.014 compared to 29 cycles; p = 0.011 compared to 30 cycles). Therefore, the Amplicon RX method should be widely adopted in forensic laboratories to enhance the analysis of extremely low-template and compromised samples. These findings highlight the potential of the amplicon RX post-PCR clean-up kit to improve trace DNA analysis in forensic casework. Further research is recommended to validate these results and explore its broader application in forensic DNA analysis, particularly in complex DNA mixtures and extremely low-template samples.