Automating direct-to-PCR for disaster victim identification

Abstract

ABSTRACTDirect-to-PCR methodology adds samples directly to PCR tubes offering gains in efficiency and sensitivity. The approach has been applied to a variety of biological sources including blood, saliva, tissue, hair and nail. We added various preservative solutions to a range of biological samples to leech DNA into solution, whilst preserving at room temperature. Tubes containing ‘free DNA’ then followed automated workflows for amplification and capillary electrophoresis. Routine FASS-automated workflows (including DNA extraction and quantification) were compared with published direct-to-PCR methodology and automated amplification of an aliquot of preservative solution. Applying preservative solutions to ~30-year-old blood stains stored at room temperature resulted in recovery of a larger quantity of DNA and more alleles (using PowerPlex 21) when compared with routine automated typing. Trials were extended to blood, saliva, hair and nail, mimicking ante-mortem samples collected in a disaster victim identification effort. Despite slightly lower allelic recovery, the faster processing times, lower costs and storage potential offers advantages for the processing of ante-mortem samples.