This document reports on a study to evaluate Illumina’s MiSeq FGx next generation sequencing platform.
This report evaluates Illumina’s MiSeq FGx NGS (next generation sequencing) Platform, which aims to address and mitigate the limitations of current DNA sequencing techniques. NGS technology adds sequencing information and has a nearly unlimited capacity for additional short tandem repeats (STRs) as well as single nucleotide polymorphisms (SNPs) that could improve individual identification, mixture deconvolution, and hereditary analysis. These upgrades could enhance individual identification and have potential for significant improvements in throughput at lower costs. Results verified concordance through current capillary electrophoresis (CE) methods. The authors note that read numbers varied tremendously within samples for specific loci, revealing an imbalance of the multiplex reaction kit, and samples with low DNA inputs had lower read numbers leading to less reliable results. They conclude that DNA input had the strongest effect on the outcome, however full STR profiles were occasionally obtained from as little DNA input as 50 pg. The authors also note genotype errors primarily occurred due to dropouts and could only be detected by comparison to repeatedly run samples; no errors were found due to contamination caused by pooling or sample handling, and spurious alleles were found in Y-STRs on a few occasions; and degraded and challenging samples resulted in almost full profiles.