Objective: To explore the cause for allelic drop-out at short tandem repeat (STR) loci upon paternity testing with a PowerPlex® 16 kit.
Methods: A total of 10 642 DNA confirmed paternity testing cases (18 314 parent/child allelic transfers) were analyzed with the PowerPlex® 16 kit. Samples suspected for having allelic drop-out were verified with an Identifiler™ kit and/or locus-specific singleplex amplification systems. PCR products of null alleles were separated and directly sequenced.
Results: Eight cases of allelic drop-out were found. The overall rate of null allele in the PowerPlex® 16 system was 0.437 × 10(-3). DNA sequencing has confirmed single base variations within the binding region of published primers, in which 4 cases involved the D18S51 locus (2 cases with G>A transitions at 79 bp upstream of the repeats, 1 case with G>T transversion at 162 bp downstream of the repeats and 1 case with G>C transversion at 74 bp upstream of the repeats), 2 cases involved the D21S11 locus (1 case with C>A transversion at 17 bp upstream of the repeats and 1 case with A>G transition at 12 bp upstream of the repeats). One case involved the FGA locus (1 case with G>A transition at 142 bp downstream of the repeats) and 1 case involved TPOX locus (1 case with G>A transition at 198 bp downstream of the repeats).
Conclusion: Base variation in the primer binding region may cause failed PCR and result in null allele reports. Alternative primer sets should be used to verify the suspected allelic drop-out. Attention should be paid to this during paternity testing and data exchange for personal identification.