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. 2008 Mar;122(2):135-42.
doi: 10.1007/s00414-007-0182-6. Epub 2007 Jun 20.

Stable RNA Markers for Identification of Blood and Saliva Stains Revealed From Whole Genome Expression Analysis of Time-Wise Degraded Samples

Free PMC article

Stable RNA Markers for Identification of Blood and Saliva Stains Revealed From Whole Genome Expression Analysis of Time-Wise Degraded Samples

Dmitry Zubakov et al. Int J Legal Med. .
Free PMC article


Human body fluids such as blood and saliva represent the most common source of biological material found at a crime scene. Reliable tissue identification in forensic science can reveal significant insights into crime scene reconstruction and can thus contribute toward solving crimes. Limitations of existing presumptive tests for body fluid identification in forensics, which are usually based on chemoluminescence or protein analysis, are expected to be overcome by RNA-based methods, provided that stable RNA markers with tissue-specific expression patterns are available. To generate sets of stable RNA markers for reliable identification of blood and saliva stains we (1) performed whole-genome gene expression analyses on a series of time-wise degraded blood and saliva stain samples using the Affymetrix U133 plus2 GeneChip, (2) consulted expression databases to obtain additional information on tissue specificity, and (3) confirmed expression patterns of the most promising candidate genes by quantitative real-time polymerase chain reaction including additional forensically relevant tissues such as semen and vaginal secretion. Overall, we identified nine stable mRNA markers for blood and five stable mRNA markers for saliva detection showing tissue-specific expression signals in stains aged up to 180 days of age, expectedly older. Although, all of the markers were able to differentiate blood/saliva from semen samples, none of them could differentiate vaginal secretion because of the complex nature of vaginal secretion and the biological similarity of buccal and vaginal mucosa. We propose the use of these 14 stable mRNA markers for identification of blood and saliva stains in future forensic practice.


Fig. 1
Fig. 1
a, b RT-PCR results for blood-targeted genes in blood and saliva stains. c RT-PCR results for saliva-targeted genes in saliva and blood stains. Genes were selected based on expression microarray results and GNF SymAtlas database. Expression values for each time point were averaged across three male and three female RNA samples; no gender-specific expression differences were detected (t test p < 0.05). B indicates blood; S indicates saliva; samples were processed after complete drying of blood and saliva at 0, 21, 57, and 180 days, respectively
Fig. 2
Fig. 2
a RT-PCR for saliva and blood-targeted genes in semen stains. b RT-PCR for saliva and blood targeted in vaginal secretion stains. Delta Ct (dCt) values were calculated as follows: dCt = Ct (candidate gene) − Ct (endogenous control, GAPDH gene). Low dCt values correspond to high expression level of the specific mRNA. Gray bars correspond to the samples from target tissues for selected genes (either blood or saliva); black bars correspond to samples from nontarget tissues (either vaginal secretion or semen). Dotted bars represent the cases were amplification was not detected after 50 cycles, in this case, the expression values were arbitrary set to Ct value of 25 (plot maximum)

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