Faculty of Health Sciences & Medicine

Future Research Projects

Improved Detection of DNA in Forensic Samples

Supervisor

Prof Angela Van Daal
Professor of Forensic Sciences

BACKGROUND

Current forensic DNA typing methods utilise between 9 and 16 STR markers to generate an individual profile.  A full profile is usually obtained from evidence and reference samples such as blood, buccal swabs and semen, due to the relatively large amounts of intact DNA.  However in circumstances in which DNA is in very low amounts and/or is highly degraded, profiles are difficult to obtain. The occurrence of several mass fatalities in the last decade has necessitated the rapid improvement of DNA techniques used in the identification of highly degraded human remains. Forensic scientists have conventionally relied on mitochondrial DNA (mtDNA) to generate a victim profile from these very difficult samples.   Although a very reliable means of identification, the discriminatory power offered by a mtDNA profile is relatively low compared to that generated by analysing genomic DNA (gDNA).  Therefore, scientists within the field are increasingly focusing on analysing gDNA from these highly degraded samples.  This requires methods to repair damaged DNA yield from a sample and more effective amplification and detection methods. 

AIMS OF THE PROJECT

This project will investigate the advantage of single nucleotide polymorphisms (SNPs) as an alternate method of identification from highly degraded bone and teeth.  SNPs are single nucleotide differences and therefore produce very short amplicons (the length of two flanking primers), making them excellent targets for analysis of extremely fragmented DNA. Preliminary data from analysis of extremely degraded samples from the World Trade Centre attacks have demonstrated the potential of SNP markers.

This project will also investigate various techniques by which the fragmented genome could be reassembled prior to amplification. This would significantly improve the likelihood of gaining a full DNA profile.

METHODS

Optimisation of downstream assays will be performed using artificially degraded DNA.  Mock case samples will be generated and exposed to various environmental conditions, burial or incineration.  A number of DNA repair methods  will be investigated for their effectiveness in repairing lesions commonly present in degraded samples, namely abasic sites, cross-links, nicks, single and double stranded breaks. An example of a method that will be investigated uses CircLigaseTM, which is an enzyme that catalyses circularization of small single stranded DNA templates.  These circular products could then become templates for rolling circle amplification methods, rescuing the highly fragmented DNA found in degraded samples.

Both PCR and whole genomic amplification (WGA) methods will be investigated. WGA is an in vitro method used to amplify large fragments of DNA. The multiple displacement amplification (MDA) process is able to amplify samples by up to 100,000 fold from a minute starting template.  The ability of MDA in conjuction with various pre-treatment protocols (eg. CircligaseTM) to accurately amplify highly degraded DNA will be investigated.