mtDNA investigations after differential lysis

In order to gain information regarding the ethnic origin of an unknown offender in a murder case it was necessary to sequence the HV1 and 2 regions of the mitochondrial DNA (mtDNA). The only evidentiary material that could be linked to the suspect was DNA, extracted from spermatozoa after differential lysis. The observed mtDNA sequences were identical to the sequences of the victim. Therefore, we had to check if this was a coincidence or the result of a technical limitation of the testing procedure. Subsequently, we performed a systematic study. In cases with complete separation of sperm and female cells it wasn't possible to obtain a mtDNA sequence for the sperm fraction. This phenomena is based on the loss of the sperm's flagellum and mid-piece during the first lysis step and a concomitant loss of the sperm's mitochondria. In our murder case, a minor carry-over of female cells to the sperm fraction was responsible for the sequencing result.