Extraction of Single-Copy Nuclear DNA from Forensic Specimens with a Variety of Postmortem Histories

Abstract

Specimens of human bone, teeth and dried blood spots from 3 months to 91 years old, with a variety of postmortem histories, were used in a comparative study of recovery of single-copy nuclear DNA sequences from forensic material. Sequences of the amelogenin and HLA-DPB1 genes were chosen for their value in sexing and identification. Sequences of the mitochondrial non-coding region V were also amplified to compare the recovery of mitochondrial and single-copy nuclear DNA. A variation of the silica method for DNA extraction was refined for application to the forensic specimens in this sample. Single-copy nuclear DNA was amplified from 100% of recent postoperative bone specimens (n = 6), 80% of forensic teeth and bone specimens (n = 10), 78% of recently extracted teeth (n = 18), 78% of exhumed bone up to 91 years old (n = 37) and 69% of 15 year old bone specimens fixed in 10% formalin (n = 20). Amelogenin sexing was correct in 85% of cases (n = 74) in which the sex of the donor had been recorded. There was no correlation between the age of the specimen and the extent of DNA preservation.

Molecular identification of human tuberculosis in recent and historic bone tissue samples: The role of molecular techniques for the study of historic tuberculosis

“…One gram of the pulverized material was then incubated with 2 ml of 0.5 M EDTA-solution containing proteinase K (0.25 mg/ml) at room temperature for 2 days on a rotatory mixer (Boom et al, 1990). Following centrifugation for 15 min at 3,000g, 0.5 ml of the supernatant was removed, and 1 ml guanidine isothiocyanate solution and diatomaceous earth (Evison et al, 1997) were added. After incubation on a rotatory mixer for another 2 hr, the diatomaceous earth was pelleted by centrifugation and washed twice with 70% ethanol and once with acetone.…”

Molecular analysis of skeletal tuberculosis in an ancient Egyptian population

“…A sample of the pulverised material (1 g) was incubated with 2 ml of 0.5 M EDTA solution containing proteinase K 0.25 mg=ml at room temperature for 2 days on a rotatory mixer [15]. Following centrifugation for 15 min at 3000 g, 0.5 ml of the supernate was removed and 1 ml of guanidine isothiocyanate solution and diatomaceous earth [16] were added. After incubation on a rotatory mixer for another 2 h, the diatomaceous earth was pelleted by centrifugation and washed twice with ethanol 70% and once with acetone.…”

Molecular evidence for different stages of tuberculosis in ancient bone samples from Hungary

“…Afterward samples were taken from the inner part of the bones and pulverized with a mixer mill (Retsch MM200, Haan, Germany). One gram of the pulverized material was then incubated with 2 mL of 0.5 M EDTA-solution containing proteinase K (0.25 mg/mL) at room temperature for 2 days on a rotatory mixer (Evison et al, 1997). After centrifugation for 15 min at 3,000 g, 0.5 mL of the supernatant was removed, and 1 mL guanidine isothiocyanate solution and diatomaceous earth (Boom et al, 1990) were added.…”