A Comprehensive Evaluation of a new Direct Amplification System (PowerPlex® 18D) in Forensic DNA Profiling

Parish-Fisher, Casie (2016) A Comprehensive Evaluation of a new Direct Amplification System (PowerPlex® 18D) in Forensic DNA Profiling. Doctoral thesis, University of Central Lancashire.

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Short tandem repeat typing is the primary method of DNA identification used in the field of forensic science. Over the past several years the need to improve on this method has moved to the forefront of research. Due to the increasing number of criminal cases and the substantial backlogs most laboratories are facing, it is vital to evaluate methods which can produce quality DNA profiles in a fast and reliable manner. Direct amplification, also referred to as direct PCR, is one alternative method that has been proposed to address this issue. Direct amplification allows for the generating of DNA profiles without using the DNA isolation process. While direct PCR would reduce processing time and resources, it is unknown if this technique would be able to generate a robust full or partial profile from samples which could be collected from scenes of crime. Often crime scene personnel must use visualization techniques, either in powder or chemical form, in order to see and collect biological evidence for submission to a crime laboratory. In order to evaluate if direct PCR is a feasible solution a comparative study between a direct PCR kit and standard DNA profiling practices was undertaken using mock crime scene type samples. Samples of this nature include surfaces which have been exposed to fingerprint powders and whole blood which has been chemically enhanced for visualization. PowerPlex® 18D, a direct amplification system, and PowerPlex® 16HS, an extraction-based method, were used to produce the profiles. An assessment of the kits aimed to critically evaluate and compare how the direct
amplification kit performs on samples which have been exposed to powder and chemical processing for visual enhancement. This will be done by reviewing two types of samples; epithelial cells which have been exposed the fingerprint powders (black, magnetic and white) and whole blood which has been exposed to chemicals (luecocrystal violet, amido black and ninhydrin). Samples subjected to direct amplification using PowerPlex® 18D generated DNA profiles with greater peak heights when compared to the extraction- based method. The peak balances for heterozygous loci were also higher and more full profiles were generated with direct amplification than with the extraction method. The amount of DNA retrieved from each substrate also varied even though the same amounts of starting material were deposited, proving that the type of substrate can affect the retrieval of DNA.

Epithelial cell samples were most successful when processed with white powder. Magnetic powder samples also yielded a positive result when using direct amplification which was not expected as in previous data magnetic powder samples have not been successful. Whole blood samples which were processed with amido black produced profiles with lower overall peak heights when compared to the two other chemical processes. This could be attributed to the rinse step which is required when working with amido black. Ninhydrin was the most successful of the chemicals in generating full, good quality profiles.

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