title: Organic extraction of bone lysates improves DNA purification with silica beads.
abstract: In our standard protocol for DNA extraction from skeletal remains of unidentified bodies, bone lysates resulting from decalcification and Proteinase K treatment were purified with the DNA IQ™ Casework Pro Kit for Maxwell®16 automate (Promega, WI). Despite its success in the majority of cases, the DNA purification with paramagnetic silica beads failed in some challenging samples. This failure in DNA recovery was often associated with filter clogging during the required volume reduction of the lysate to enable loading on the automate. Two modifications to the standard method were tested for a more efficient filtering and purification. Adding collagenase to the lysate reduced the filter lead time but did not enhance DNA yield, while organic extraction of the crude lysate solved the filter clogging and resulted in successful DNA purification. The modified method in which a phenol treated lysate was loaded on the automate resulted in successful STR-profiling of the skeletal remains of all 13 unidentified bodies tested, which showed a wide variety in post mortem interval and preservation conditions. The variation in DNA yield between the 28 samples tested showed the importance of bone type selection and multiple sampling in successful STR-profiling of skeletal remains. Despite the disadvantages inherent to phenol, the organic extraction of crude bone lysates enhanced the efficiency of DNA purification with paramagnetic silica beads. The combined method of organic extraction and purification with silica beads resulted in STR-profiling of challenging bone samples.
authors.affiliation: National Institute of Criminalistics and Criminology, Brussels, Belgium. Electronic address: stijn.desmyter@just.fgov.be.
authors.forename: Stijn
authors.initials: S
authors.lastname: Desmyter
authors.affiliation: National Institute of Criminalistics and Criminology, Brussels, Belgium.
authors.forename: Greet
authors.initials: G
authors.lastname: De Cock
authors.affiliation: National Institute of Criminalistics and Criminology, Brussels, Belgium.
authors.forename: Sabine
authors.initials: S
authors.lastname: Moulin
authors.affiliation: National Institute of Criminalistics and Criminology, Brussels, Belgium.
authors.forename: Fabrice
authors.initials: F
authors.lastname: Noël
mesh_terms: D001842:Bone and Bones / Q000737:chemistry*; D005591:Chemical Fractionation / Q000379:methods; D004247:DNA / Q000302:isolation & purification*; D016172:DNA Fingerprinting; D003648:Decalcification Technique; D019286:Endopeptidase K / Q000494:pharmacology; D006801:Humans; D060328:Magnetic Phenomena; D018895:Microsatellite Repeats; D008863:Microspheres; D012822:Silicon Dioxide / Q000737:chemistry*
chemical_list: D012822:Silicon Dioxide; D004247:DNA; D019286:Endopeptidase K
keywords: Bone; DNA; Organic extraction; Paramagnetic beads; Silica