Currently, forensic identification techniques used in cases involving pets at crime scenes often rely on shed hair analysis as it might relate to the victim or suspect. However, shed hair analysis can be difficult since they often do not contain the bulbous root that contains nuclear or mitochondrial DNA. There are some short tandem repeat (STR) multiplex kits for animals that are already in use. However, most of these STR kits, like the Meowplex used for identifying cats, focuses on one species identification. While autosomal STRs may provide a means for specific identification of an animal, mitochondrial DNA degrades slower than nuclear DNA and is much more plentiful within cells; therefore, this type of DNA can often be useful when evidence from an unknown source is collected at a crime scene. When hair, tissue, or blood are found at a crime scene and no animal is present, a method to identify the unknown species as the source of the evidence is needed before moving to STR analysis.
The objective of this project will use mitochondrial DNA markers to create a reference ladder and design a multiplex kit that can simultaneously detect species of dog, cat, ferret, and rabbit, common domestic pets that may be associated with a crime scene. A labeled universal forward primer and species-specific reverse primers will be used for simultaneous amplification of four species in one multiplex. Successful single-plex amplification of dog and cat mitochondrial DNA has been done. The individual species' products are mixed together and used as a reference ladder. Any unknown samples can quickly be identified when aligned with the known reference ladder on the genetic analyzer. The next step is to incorporate dog, cat, ferret, and rabbit primers into the multiplex and validate it.