SD42: Effect of preservation, time and temperature on DNA recovery, quantification, and amplification on blow fly specimens (Diptera: Calliphoridae)

Correct blow fly (Diptera: Calliphoridae) species identification is a fundamental prerequisite of any minimum time of death investigation using insect evidence. To successfully identify preserved samples, standard operating procedures must be followed, using the correct storage conditions and preservative. In situations where specimens are stored incorrectly, or poorly preserved/damaged, genomic and mitochondrial DNA is relied upon to amplify, sequence, and analyze a barcoding region for species identity. Whilst it is commonly agreed in the forensic entomology community that 70-80% ethanol after hot water killing of larval specimens should be used, the effect of temperature, time and preservative has not been extensively examined in relation to the success rate of retrieving and sequencing DNA. This project aimed to examine the effect that different preservation treatments and time had on the success rate of recovering DNA for amplification and species identification. Larvae (n=12) of Calliphora latifrons (Diptera: Calliphoridae) were sampled and underwent a set number of treatments. Four preservatives, time in preservation and temperature were examined. DNA was extracted and quantified using QuBit fluorimetry. An approximate 600 base pair region of the cytochrome oxidase I was amplified to determine successful amplification of the species barcoding region. Statistical analysis was applied to DNA quantification data and found that preservatives had a significant effect on the recovery and success rate of amplification. Future work is expected to examine the effect of longer time intervals, more preservatives, and different temperatures.