The development of a DNA analysis system for pollen

Abstract

The use of pollen grains for forensic evidence was established in 1959 and has since been used successfully in many cases. The traditional method of pollen analysis is microscopy where the pollen grains are identified by the distinct patterns on the pollen wall. This is time consuming and requires a trained and experienced palynologist. This thesis aimed to develop a DNA analysis system for pollen grains extracted from soil that could be applied to forensic samples in casework. A DNA analysis technique would be more beneficial than microscopy because it is quick and simple, it does not require in depth knowledge of the pollen types and reduces subjectivity associated with human judgment. The DNA analysis technique, tRFLP, was tested using the plant material from eight different species. The results were analysed using a Euclidean distance measure to compare samples. The analysis showed that the tRFLP technique had good reproducibility and that there was high discrimination between different species. A technique using glass bead maceration was found to be suitable for extracting amplifiable DNA from pollen grains. Analysis of the DNA from five pollen species also indicated good reproducibility and discrimination between species. The method of using zinc bromide to remove pollen grains from soil was not successful and an alternative method using sucrose is suggested. The results from the work undertaken for this thesis established that the tRFLP technique was a reliable and reproducible technique that can provide considerable discriminating power between samples. It will be suitable for application to forensic casework samples but requires further research and validation.