Double incompatibility at human alpha fibrinogen and penta E loci in paternity testing

We present a paternity testing case in which a double incompatibility was found for two short tandem repeat (STR) markers, human fibrinogen alpha (FGA) and Penta E. Analysis of the trio (mother, father, and daughter) included the amplification with a battery of 15 autosomal short tandem repeats (STR) by using a commercially available PowerPlex 16 System kit, and the detection with an ultraviolet-automatic sequencer. The biological paternity was confirmed with 12 additional markers. Reanalysis of the trio for the same markers with different primers was carried out by using an infrared automated sequencer and infrared-fluorescent primers. High paternity index confirmed that the observed inconsistencies were due to a double mutation, which was confirmed by sequence analysis at FGA and Penta E loci. Amplification and detection results obtained by the infrared-protocol showed consistent results with those obtained by ultraviolet-protocol and a commercially available kit. This has been our first case of double mutation at FGA and Penta E in a paternity testing. The use of our approach, based on two amplification and detection formats and on the sequence analysis, confirmed the observed meiotic paternal mutations.