The Investigator ESSplex SE QS Kit (Qiagen) is a next-generation polymerase chain reaction (PCR) kit that, in 60 min, amplifies 17 Short Tandem Repeat (STR) markers, including the five European Standard Set (ESS) loci (D10S1248, D12S391, D1S1656, D22S1045, D2S441), the SE33 marker, and the locus Amelogenin for sex determination. Two quality sensors (QS1 and QS2) are also co-amplified to check PCR performance. Since forensic laboratories carry out hundreds of DNA typings annually, we verified the kit’s performance using half reaction volumes with the aim of improving the number of samples that may be amplified with a single kit and consequently reducing laboratory costs. In the present study, intended as a technical note rather than internal validation, some control samples (oral swabs) with known DNA profiles and 40 real casework samples were analyzed. We observed that reducing the total reaction volume, while keeping all component ratios unaltered, yields DNA profiles comparable to those obtained using standard reaction volumes and with allele peaks higher than those with regular volumes. Using half volumes for PCR amplification enables the analysis of a larger number of samples compared to the standard protocol, thereby reducing laboratory costs without compromising the quality of the analysis.

Background: the Investigator ESSplex SE QS Kit (Qiagen) is a next generation PCR kit that, in 60 minutes, amplifies 17 STRs markers, including the 5 European Standard Set (ESS) loci, the SE33 marker and the locus Amelogenin for sex determination. Two Quality Sensors (QS1 and QS2) are co-amplified to check PCR performance. Since forensic laboratories carry out annually hundreds of DNA typing, we verified the kit performance using half reaction volumes with the aim to improve the number of samples that may amplified with a kit, reducing consequently the laboratory costs. Methods: in the study, that is intended as a technical note and not an internal validation, were analyzed control samples (oral swabs) with known DNA profiles and 40 real caseworks samples. Results: we observed that reducing the total reaction volume, while keeping unaltered all components ratio’s, yields DNA profiles comparable to the ones obtained using standard reaction volumes and with alleles peaks higher than with standard volumes.. Conclusions: the use of half volumes for PCR amplification allows the analysis of a wider number of samples than the standard protocol, reducing laboratory costs while maintaining unaltered the quality level of the analysis.

Bones and teeth often represent the only sources of DNA available for identifying human remains. DNA in bones and teeth is generally better preserved than that in soft tissues because of the presence of hard connective tissue with a high level of calcium. Because of the extensive mineralisation, the choice of an efficient DNA extraction procedure is important to minimise the sampling of a high level of minerals and to remove polymerase chain reaction (PCR) inhibitors. Some protocols are available for DNA extraction from bones and teeth as part of the Qiagen EZ1 DNA Investigator Kit using the EZ1 Advanced XL automated purification platform. To improve the efficiency of DNA extraction from skeletal remains, the present study focuses on a modification to these already available protocols. In this study, different bones and teeth collected between 1 and 50 years after death were subjected to DNA extraction using the standard EZ1 protocol, a supplementary protocol, and a modified protocol. The modified approach included a decalcification step, whereas the Qiagen protocols worked directly on non-decalcified powder. In all three procedures, 150 mg samples were used for DNA extraction. We evaluated the quantity of DNA recovered from samples, the presence of any PCR inhibitors co-extracted, the level of DNA degradation, the quality of short tandem repeat (STR) profiles, and the reproducibility of the modified procedure. When compared with the other protocols, the modified protocol resulted in the best recovery of DNA that was free of PCR inhibitors. Additionally, the STR profiles were reliable and of high quality. In our opinion, the decalcification step increases DNA recovery by softening tissues, which allows lysis solutions to act more effectively. Furthermore, the use of two lysis solutions and the variation added to the EZ1 purification step allow for DNA recovery with quality and quantity superior to those of the previously available Qiagen-based protocols. These findings may be helpful solutions to the problems commonly encountered when dealing with difficult samples, such as bones and teeth.

Identifiler™ is the most recent kit from Applied Biosystems. It uses a five-dye fluorescent system and so coamplifies the repeat regions of 15 STR loci and a segment of the X–Y homologous gene Amelogenin. In the present study, we analyzed allelic and genotypic distribution of STRs loci in three populations from Southern Italy (Calabria): Reggio Calabria, Catanzaro, and Cosenza.