Implementation of RNA profiling in forensic casework

An essential aspect for forensic methods is the prevention of cognitive (confirmation, expectation or motivational) bias. While implementing RNA profiling in casework, we developed a stepwise procedure for unbiased assessment in which: (1) the RNA researcher who generates DNA/RNA fractions and performs RNA profiling, remains uninformed about the context of the case and (2) presents RNA profiling results that are derived by clear guidelines in a results table that uses six different scoring categories, (3) the DNA fractions are processed and analysed by DNA analysts following the standard routine after which (4) reporting officers interpret the DNA profiles and establish the relation to the RNA results which is succeeded by (5) collating all generated results in the case and formulating conclusions in expert reports. The scoring guidelines and results table have a general purpose and can apply to any RNA multiplex. This procedure was applied in a comparative study encompassing seven mock cases designed to be especially interesting for body fluid identification by RNA profiling. Samples were prepared in duplicates and subjected to either presumptive testing combined with standard DNA typing or RNA/DNA co-extraction followed by RNA and DNA profiling. For all cases, the results from presumptive testing and RNA profiling agreed to the level of details the tests can give and concordant DNA results were obtained. RNA profiling was especially useful when (1) menstrual secretion and peripheral blood needed to be distinguished, (2) presence of vaginal mucosa was questioned or (3) presence of skin cells was informative. For forensic reports, we propose to use sets of hypotheses evaluated by the conclusions obtained with DNA and RNA analyses.