Introduction

Touched objects and fingerprints have been used in forensic biology to generate DNA profiles for over 10 years

Methods for cellular origin identification

1. Phadebas test saliva

Methods for cellular origin identification

2. ABAcard HemaTrace method blood

Methods for cellular origin identification

3. Prostate specific antigen detection of semen

Problem:

Current DNA profiling method----inefficient!

Solution:

mRNA

Short tandem repeat (STR) analysis

evaluates specific STR regions found on nuclear DNA. Method for isolating DNA samples
Probability of misidentification can be as high as 2 in 1 billion or greater

The messenger RNA (mRNA)

Translates genetic code of DNA Provides necessary specificity, sensitivity and automation.

mRNA markers
Identifiable in the differences expression patterns of transciptomes. A result of mRNA profiling Substitute to DNA profiling results in

Our outer epithelium

Continually being renewed
Complete renewal ever 40-56 days

Progressively keratinized
Aided by different proteins
Keratin, filaggrin, involucrin and loricrin

Candidate genes
CALML5, CDSN, DCD, DSC1, FLG, KRT1, K RT2A,KRT10, LOR and SCGB2A2 KRT9

mRNA markers
KRT9 CDSN LOR

KRT9 gene
Type I keratin Expressed in the suprabasal cells of the epidermis Specifically expressed in palm and soles of the feet

CDSN
52- to 56-kDa basic glycoprotein Encodes a protein found in corneodesmosomes

LOR
expressed in the granular layer of the epidermis 80% of the total protein mass of the cornified envelope

Specificity of skin-targeted mRNA markers

Fig. 1 Ct values for three skin-targeted mRNA markers CDSN, KRT9 and LOR in various skin samples and across forensically relevant body fluids:

Specificity of skin-targeted mRNA markers

Fig. 1 Ct values for three skin-targeted mRNA markers CDSN, KRT9 and LOR in various skin samples and across forensically relevant body fluids:

Exceptions in the specificity of skin-targeted mRNA markers

Fig. 1 Ct values for three skin-targeted mRNA markers CDSN, KRT9 and LOR in various skin samples and across forensically relevant body fluids:

Exceptions in the specificity of skin-targeted mRNA markers

Non-skin

Skin

Fig. 1 Ct values for three skin-targeted mRNA markers CDSN, KRT9 and LOR in various skin samples and across forensically relevant body fluids:

Sensitivity of skin-targeted mRNA markers

3 collection methods tested 1. wet/dry cotton swabbing 2. wet/dry swabbing with flocked swabs 3. Pinpoint Slide RNA isolation System II kit

Sensitivity of skin-targeted mRNA markers

Pinpoint method
Allowed reproducible detection of skin-targeted mRNA markers Used for skin cell collection due to an expected minute amount of available material

Sensitivity of skin-targeted mRNA markers

Fig. 2 Detection sensitivity of qPCR assays for three skin-targeted mRNA markers CDSN, KRT9 and LOR, with full (n=5), half (n=5) and quarter (n=10) thumbprints.

Sensitivity of skin-targeted mRNA markers

Fig. 2 Detection sensitivity of qPCR assays for three skin-targeted mRNA markers CDSN, KRT9 and LOR, with full (n=5), half (n=5) and quarter (n=10) thumbprints.

Time wise stability of skin-targeted mRNA markers

Fig. 4 Time-wise stability of three skin-targeted mRNA markers CDSN, KRT9 and LOR after 6.5 months of thumbprint storage

Time wise stability of skin-targeted mRNA markers

Fig. 4 Time-wise stability of three skin-targeted mRNA markers CDSN, KRT9 and LOR after 6.5 months of thumbprint storage

The mRNA markers

KRT9 CDSN LOR