Novel extraction chemistry and alternative amplification strategies for use with rootless hair shafts

Abstract Rootless hair shafts are often considered unsuitable for STR genotyping due to the known high failure rate. The same samples can be reliably processed with mitochondrial sequencing. However, the minimal discriminatory power of widely implemented control region mitochondrial sequencing techniques limits its utility in some forensic casework. In this research, multiple variables were tested to provide information on rootless hair shaft sample genotyping success. Results showed external decontamination procedures decreased drop‐in alleles but also greatly reduced profile recovery. The novel InnoXtract ™ chemistry was comparable to automated EZ1 DNA Investigator extraction. With thoroughly decontaminated hairs, InnoTyper ® 21 amplification generated random match probabilities higher than STR chemistry in 71.875% of samples and 18.75% of samples benefitted from the use of InnoTyper ® 21 amplification compared with estimated mtDNA profile rarity. Compared with the capillary electrophoresis‐based amplification chemistries tested, the ForenSeq™ DNA Signature Prep chemistry paired with massively parallel sequencing was the most discriminatory amplification strategy tested.