Challenges in mRNA-based time since deposition estimation: A multiplex MPS primer panel investigation

Area of Science:

• Forensic science
• Molecular biology
• Genetics

Background:

• Estimating the time since deposition (TsD) of biological stains is critical for crime scene investigations.
• Current methods for TsD estimation are limited, necessitating the development of new approaches.

Purpose of the Study:

• To investigate mRNA degradation patterns in four body fluids (blood, semen, menstrual blood, vaginal secretion) for TsD estimation.
• To assess the utility of a custom mRNA MPS primer panel for analyzing degradation patterns.

Main Methods:

• Utilized a custom mRNA MPS primer panel designed with candidate markers identified via RNA-Seq.
• Analyzed mRNA degradation based on individual trends, intra-transcript stability (5' vs. 3' ends), and inter-transcript stability.

Main Results:

• Observed significant inter-individual variability in mRNA degradation, particularly for low-abundance transcripts.
• Identified some mRNA markers whose degradation patterns correlated with TsD.
• Encountered inconsistencies in degradation patterns, indicating challenges in primer panel optimization, RNA extraction, and reproducibility.

Conclusions:

• mRNA degradation shows potential for forensic TsD estimation.
• Forensic application requires careful selection and validation of markers across diverse datasets to address biological and methodological variability.