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Abstract
The efficient extraction of DNA from challenging samples, such as bones, is critical for the success of downstream genotyping analysis in molecular genetic disciplines. Even though the ancient DNA community has developed several protocols targeting small DNA fragments that are typically present in decomposed or old specimens, only recently forensic geneticists have started to adopt those protocols. Here, we compare an ancient DNA extraction protocol (Dabney) with a bone extraction method (Loreille) typically used in forensics. Real-time quantitative PCR and forensically representative typing methods including fragment size analysis and sequencing were used to assess protocol performance. We used four bone samples of different age in replicates to study the effects of both extraction methods. Our results confirm Loreille’s overall increased gain of DNA when enough tissue is available and Dabney’s improved efficiency for retrieving shorter DNA fragments that is beneficial when highly degraded DNA is present. The results suggest that the choice of extraction method needs to be based on available sample, degradation state, and targeted genotyping method. We modified the Dabney protocol by pooling parallel lysates prior to purification to study gain and performance in single tube typing assays and found that up to six parallel lysates lead to an almost linear gain of extracted DNA. These data are promising for further forensic investigations as the adapted Dabney protocol combines increased sensitivity for degraded DNA with necessary total DNA amount for forensic applications.
Highlights
This article is an open access articleDNA extraction is the first and perhaps most important step in any DNA analysis workflow
In the fields of ancient DNA and forensic genetics, where the samples under investigation are often highly degraded and DNA is present in low copy number, highly efficient DNA extraction methods for skeletal remains are elemental
DNA preserved in skeletal remains and sediments from up to hundreds of thousands of years [1,2,3,4], the continuous research on new and improved extraction methods indicates the boundaries for efficient DNA isolation and recovery are still to be defined [5,6,7,8,9,10,11,12,13,14,15]
Summary
This article is an open access articleDNA extraction is the first and perhaps most important step in any DNA analysis workflow. In the fields of ancient DNA and forensic genetics, where the samples under investigation are often highly degraded and DNA is present in low copy number, highly efficient DNA extraction methods for skeletal remains are elemental. DNA preserved in skeletal remains and sediments from up to hundreds of thousands of years [1,2,3,4], the continuous research on new and improved extraction methods indicates the boundaries for efficient DNA isolation and recovery are still to be defined [5,6,7,8,9,10,11,12,13,14,15]. The DNA is washed with an ethanol-based solution to minimize inhibitor carry-over and eluted in a low concentrated salt buffer
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