The SE33 locus is one of the most polymorphic markers used in human identification. However, it also possesses multiple microvariants both within the repeat and in the flanking regions. Such flanking region mutations can generate discordant allele calls between multiplexes using different primer pairs. Classically such discordance is due to a mutation within a primer binding site. However, a discordance observed in samples of primarily West African descent between the PowerPlex ESI 17 System and other commercial STR multiplexes does not fall in this category. The root cause is a set of up to four separate single nucleotide polymorphisms (SNPs) residing within the PowerPlex ESI 17 SE33 amplicon, which are capable of disrupting a stem-loop structure present in the wild-type sequence. These mutations retard the migration of the amplicon on capillary electrophoresis by almost one base relative to the wild-type amplicon. To alleviate this discordance we developed a new SE33 primer set for inclusion in the PowerPlex ESI 17 Pro System. This change allows robust amplification of DNA samples containing these SNPs without the migration shift. Master Mix and primer pairs for the remaining 15 autosomal loci and amelogenin are identical between the PowerPlex® ESI 17 Pro System and the PowerPlex ESI 17 System. While the size of the SE33 amplicons is similar in both systems, allowing use of existing PowerPlex ESI panels and bins text files, a new allelic ladder is provided with the PowerPlex ESI 17 Pro System with SE33 alleles generated using the new primer pair. A new internal lane standard is also provided with the PowerPlex ESI 17 Pro System which is based on the current CC5-labelled Internal Lane Standard 500 (ILS 500) but contains competitor oligonucleotides that prevent reannealing of the new PowerPlex ESI 17 Pro SE33 primers to the denatured SE33 amplicon following electrokinetic injection.
Citation: Forensic Science International: Genetics
Pub Type: Journals
Forensic, DNA, forensic DNA, short tandem repeat, STR, SE33, concordance, validation, miniSTR, PowerPlex