Development of a combined differential DNA/RNA co-extraction protocol and its application in forensic casework

Abstract

In forensic casework, a broad range of extraction techniques is available, and the type of case will decide on the extraction method. Generally, in sexual assault cases, either the presence of semen is questioned (foremost in victim samplings), or the presence of body fluids such as vaginal mucosa or saliva (mostly in samples taken from the suspect). To analyze the semen component, differential extraction (DE) procedures are helpful as spermatozoa DNA gets specifically extracted in a distinct fraction. To assess which body fluid is present, messenger RNA (mRNA) profiling has emerged as one the best methods especially for body fluids encountered in forensic sexual assault cases. mRNA profiling requires the use of DNA/RNA co-extraction methods. In common sexual assault cases, mRNA typing is requested whilst semen may also be present, possibly in surplus amounts. This drove the development of a combined DNA/RNA/DE extraction method that yields DNA and RNA of both the sperm fraction as well as the non-sperm fraction. For the validation of the developed method, its performance was successfully assessed against conventional extraction methods, a range of relevant body fluids and stains up to 1 month old. The developed protocol was implemented for use in casework and is now part of the regular casework workflow in our laboratory.