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Abstract
BackgroundTeeth are a valuable source of DNA for identification of fragmented and degraded human remains. While the value of dental pulp as a source of DNA is well established, the quantity and presentation of DNA in the hard dental tissues has not been extensively studied. Without this knowledge common decontamination, sampling and DNA extraction techniques may be suboptimal. Targeted sampling of specific dental tissues could maximise DNA profiling success, while minimising the need for laborious sampling protocols and DNA extraction techniques, thus improving workflows and efficiencies. We aimed to determine the location of cellular DNA in non-degraded human teeth to quantify the yield of nuclear DNA from cementum, the most accessible and easily sampled dental tissue, and to investigate the effect of a common decontamination method, treatment with sodium hypochlorite (bleach).We examined teeth histologically and subsequently quantified the yield of nuclear DNA from the cementum of 66 human third molar teeth. We also explored the effects of bleach (at varying concentrations and exposure times) on nuclear DNA within teeth, using histological and quantitative PCR methods.ResultsHistology confirmed the presence of nucleated cells within pulp and cementum, but not in dentine. Nuclear DNA yields from cementum varied substantially between individuals but all samples gave sufficient DNA (from as little as 20 mg of tissue) to produce full short tandem repeat (STR) profiles. Variation in yield between individuals was not influenced by chronological age or sex of the donor. Bleach treatment with solutions as dilute as 2.5% for as little as 1 min damaged the visible nuclear material and reduced DNA yields from cementum by an order of magnitude.ConclusionsCementum is a valuable, and easily accessible, source of nuclear DNA from teeth, and may be a preferred source where large numbers of individuals need to be sampled quickly (for example, mass disaster victim identification) without the need for specialist equipment or from diseased and degraded teeth, where pulp is absent. Indiscriminant sampling and decontamination protocols applied to the outer surface of teeth can destroy this DNA, reducing the likelihood of successful STR typing results.
Highlights
Teeth are a valuable source of DNA for identification of fragmented and degraded human remains
They were noted in accessory canals, in soft tissue inclusions, and in bone and soft tissue remnants that were present in teeth with constricted furcation areas
In three out of four teeth the cellular cementum was seen to begin approximately two-thirds up the root but on the fourth tooth it started quite close to the enamel junction
Summary
Teeth are a valuable source of DNA for identification of fragmented and degraded human remains. While the value of dental pulp as a source of DNA is well established, the quantity and presentation of DNA in the hard dental tissues has not been extensively studied Without this knowledge common decontamination, sampling and DNA extraction techniques may be suboptimal. We aimed to determine the location of cellular DNA in non-degraded human teeth to quantify the yield of nuclear DNA from cementum, the most accessible and sampled dental tissue, and to investigate the effect of a common decontamination method, treatment with sodium hypochlorite (bleach). Pulp is largely cellular and has no mineral content Both calcium and collagen have been shown to be inhibitors of polymerase chain reaction (PCR) amplification [12] and as such their coextraction needs to be minimised. EDTA is a PCR inhibitor so needs to be removed along with calcium and collagen prior to downstream processes
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