Efficiency evaluation of a DNA extraction and purification protocol on archival formalin-fixed and paraffin-embedded tissue.
Formalin-fixed and paraffin-embedded tissue (FF-PET) is an invaluable resource for retrospective molecular genetic studies, but the extraction of high-quality genomic DNA from FF-PET is still a problematic issue. Despite the range of DNA extraction methods currently in use, the association of phenol-chloroform extraction and silica-based purification protocols, reported in ancient DNA studies on archaeological bones, has, to our knowledge, not been used for DNA extraction from FF-PET yet. The present study compared the efficiency of three DNA extraction and purification protocols from two different FF-PET substrates, heart and liver, by using quantitative PCR and multiplex amplification. We showed that the method, using phenol-chloroform and the QIAamp DNA mini Kit (Qiagen), was the most effective DNA extraction and purification method and that the DNA quantity extracted from liver is statistically more important than that extracted from heart. Autosomal STR typing by multiplex amplifications gave partial allelic profiles with only small size products (less than 300 bases) amplified, suggesting that DNA extracted from FF-PET was degraded. In conclusion, the protocol presented here, previously described in studies on ancient bones, should find application in different molecular studies involving FF-PET material.
Institute of Legal Medicine, EA4438, University of Strasbourg, Strasbourg, France. audrey.farrugia@unistra.fr
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Farrugia
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Keyser
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Ludes
D002725:Chloroform; D004247:DNA / Q000302:isolation & purification*; D053836:DNA Degradation, Necrotic; D016172:DNA Fingerprinting; D005404:Fixatives; D053803:Forensic Genetics / Q000379:methods; D005557:Formaldehyde; D006801:Humans; D008099:Liver / Q000473:pathology; D009206:Myocardium / Q000473:pathology; D016612:Paraffin Embedding; D019800:Phenol; D016133:Polymerase Chain Reaction; D012997:Solvents; D020080:Tandem Repeat Sequences
D005404:Fixatives; D012997:Solvents; D005557:Formaldehyde; D019800:Phenol; D002725:Chloroform; D004247:DNA