Non-uniform phenotyping of D12S391 resolved by second generation sequencing

Area of Science:

• Forensic Science
• Genetics
• Molecular Biology

Background:

• Non-uniform phenotyping at the D12S391 locus complicates forensic DNA analysis.
• Standard STR typing kits can produce inconsistent results for complex loci.

Purpose of the Study:

• To investigate the causes of non-uniform phenotyping at the D12S391 locus.
• To characterize microvariants and off-ladder alleles at D12S391 using advanced sequencing techniques.

Main Methods:

• Utilized AmpFℓSTR NGM SElect PCR Amplification Kit and GeneMapper ID-X for initial typing.
• Employed PowerPlex ESX 17 system for confirmation.
• Conducted second-generation sequencing on samples with microvariants or off-ladder alleles.

Main Results:

• Observed non-uniform phenotyping in D12S391 locus samples.
• Identified heterozygous individuals with alleles differing by one nucleotide.
• Poor separation of alleles was noted when the short allele had more AGAT repeats than the long allele.
• Sequencing of 45 individuals revealed 30 different alleles, 16 of which were novel.

Conclusions:

• The D12S391 locus presents challenges in forensic STR analysis due to complex repeat structures and microvariants.
• Second-generation sequencing is crucial for accurately identifying and characterizing novel alleles and microvariants.
• Findings contribute to improved accuracy and reliability in forensic DNA profiling.