Massively parallel sequencing of 12 autosomal STRs in Cannabis sativa

Massively parallel sequencing (MPS) is an emerging technology in the field of forensic genetics that provides distinct advantages compared to capillary electrophoresis. This study offers a proof of concept that MPS technologies can be applied to genotype autosomal STRs in Cannabis sativa. A custom panel for MPS was designed to interrogate 12 cannabis-specific STR loci by sequence rather than size. A simple workflow was implemented to integrate the custom PCR multiplex into a workflow compatible with the Ion Plus Fragment Library Kit, Ion™ Chef, and Ion™ S5 System. For data sorting and sequence analysis, a custom configuration file was designed for STRait Razor v3 to parse and extract STR sequence data. This study represents a preliminary investigation of sequence variation for 12 autosomal STR loci in 16 cannabis samples. Full concordance was observed between the MPS and CE data. Results revealed intra-repeat variation in eight loci where the nominal or size-based allele was identical, but variances were discovered in the sequence of the flanking region. Although only a small number of cannabis samples were evaluated, this study demonstrates that more informative STR data can be obtained via MPS.