# In the trail of the crime scene dog: Processing of human DNA from outdoor samples

## TL;DR

This paper presents an optimized DNA extraction protocol for outdoor crime scene samples, significantly improving DNA recovery and STR profiling from soil and other matrices.

## Contribution

A 43-fold increase in DNA yield from outdoor samples using an optimized soil microbial DNA extraction protocol.

## Key findings

- The protocol produced high-quality STR profiles from semen in 10 mL of various soil types and matrices.
- Extracted DNA was over 95% double-stranded, suitable for library preparation methods like whole genome sequencing.
- The method successfully generated human STR profiles in 50% of casework, where traditional methods failed in 82% of those cases.

## Abstract

The growing use of crime scene dogs to detect human body fluids has led to an increased number of outdoor samples being submitted to forensic DNA laboratories. The analysis of DNA in outdoor samples such as soil presents significant challenges with present protocols due to the combination of relatively large sample volumes and co-extraction of humic substances. To address this, we have optimised a soil microbial DNA extraction protocol for the recovery of semen by incorporating enzymatic, chemical and physical lysis steps. This optimisation increased the DNA yield 43-fold. Using mock crime scene samples, the protocol provided high quality STR profiles from semen in 10 mL of different types of soil (mixed forest soil, sandy soil and park soil), organic matrix (dry leaves) and inorganic matrices (sand and gravel). The protocol also generated complete STR profiles for soil samples containing blood, saliva and cell-free DNA. However, all humic substances were not removed, causing inhibition in MPS analysis. The extracted DNA consisted of more than 95% double-stranded (ds) DNA with equivalent fragment sizes as DNA extracted using magnetic bead-based kits. The protocol is thus compatible with library preparation methods that demand dsDNA, e.g. in whole genome sequencing. The protocol has been successfully implemented in casework and used in serious crime investigations. Human STR profiles have been generated from crime scene items in 50% of the cases. In the majority of these (82%), no results were generated using swabs and traditional DNA extraction methods.