Highlights
- •Developed a multiplex digital gene expression method for body fluid identification.
- •Uses solution hybridization of color-coded NanoString® probes to 23 mRNA targets.
- •We devised a simple 5 min room temperature lysis protocol for RNA isolation.
- •Maximum likelihood algorithm to estimate body fluid quantities in a sample.
- •Use likelihood ratios to test for the presence of each body fluid in a sample.
Abstract
A DNA profile from the perpetrator does not reveal, per se, the circumstances by which it was transferred. Body fluid identification by mRNA
profiling may allow extraction of contextual ‘activity level’ information from forensic
samples. Here we describe the development of a prototype multiplex digital gene expression
(DGE) method for forensic body fluid/tissue identification based upon solution hybridization
of color-coded NanoString® probes to 23 mRNA targets. The method identifies peripheral blood, semen, saliva,
vaginal secretions, menstrual blood and skin. We showed that a simple 5 min room temperature cellular lysis protocol gave equivalent results to standard RNA
isolation from the same source material, greatly enhancing the ease-of-use of this
method in forensic sample processing.
We first describe a model for gene expression in a sample from a single body fluid
and then extend that model to mixtures of body fluids. We then describe calculation
of maximum likelihood estimates (MLEs) of body fluid quantities in a sample, and we
describe the use of likelihood ratios to test for the presence of each body fluid
in a sample. Known single source samples of blood, semen, vaginal secretions, menstrual
blood and skin all demonstrated the expected tissue-specific gene expression for at
least two of the chosen biomarkers. Saliva samples were more problematic, with their
previously identified characteristic genes exhibiting poor specificity. Nonetheless
the most specific saliva biomarker, HTN3, was expressed at a higher level in saliva
than in any of the other tissues.
Crucially, our algorithm produced zero false positives across this study's 89 unique
samples. As a preliminary indication of the ability of the method to discern admixtures
of body fluids, five mixtures were prepared. The identities of the component fluids
were evident from the gene expression profiles of four of the five mixtures. Further
optimization of the biomarker ‘CodeSet’ will be required before it can be used in
casework, particularly with respect to increasing the signal-to-noise ratio of the
saliva biomarkers. With suitable modifications, this simplified protocol with minimal
hands on requirement should facilitate routine use of mRNA profiling in casework laboratories.
Keywords
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Article info
Publication history
Published online: September 14, 2014
Accepted:
September 7,
2014
Received in revised form:
August 27,
2014
Received:
July 23,
2014
Identification
Copyright
© 2014 Elsevier Ireland Ltd. Published by Elsevier Inc. All rights reserved.
