A novel, 4-h DNA extraction method for STR typing of casework bone samples.

Bones are often found in mass grave crime scene. To increase DNA identification success rates, a highly efficient DNA extraction method should be selected. Several DNA extraction methods for human bones have been published yet never been systematically compared, and some are time-consuming or complex. As such, a quick and highly efficient DNA extraction method was developed and compared with three published methods (Hi-Flow silica-based, total demineralization (TD) and PrepFiler BTA) using 70 fresh and 22 casework bones from different body parts. The highest median DNA concentrations were obtained from developed method (135.85 ng/μL and 0.224 ng/μL for fresh and casework bones, respectively). For residual PCR inhibitors, the threshold cycle (Ct) of the internal positive control (IPC) showed that developed method and PrepFiler BTA removed most PCR inhibitors. Similarly, 95.45% of casework STR profiles obtained using the developed protocol meet the standard requirements for Australian National Criminal Investigative DNA Database (NCIDD) entry, followed by 86.35% using TD, 81.82% using PrepFiler BTA, and 45.45% using Hi-Flow. Additionally, DNA extracts from seven different bones revealed that the 1st distal phalange of the hand contained the highest DNA concentration of 338.43 ng/μL, which was three times higher than the tibia and femur. Our findings suggest that developed method was highly efficient for casework bone analysis. It significantly reduced the extraction processing time down to 4 h and is two to four times cheaper compared with other methods. In practice, both the extraction method and the bone sampling must be considered by a forensic DNA analyst to increase the chances of successful identification.