Technical note: removal of metal ion inhibition encountered during DNA extraction and amplification of copper-preserved archaeological bone using size exclusion chromatography.
A novel technique for the removal of metal ions inhibiting DNA extraction and PCR of archaeological bone extracts is presented using size exclusion chromatography. Two case studies, involving copper inhibition, demonstrate the effective removal of metal ion inhibition. Light microscopy, SEM, elemental analysis, and genetic analysis were used to demonstrate the effective removal of metal ions from samples that previously exhibited molecular inhibition. This research identifies that copper can cause inhibition of DNA polymerase during DNA amplification. The use of size exclusion chromatography as an additional purification step before DNA amplification from degraded bone samples successfully removes metal ions and other inhibitors, for the analysis of archaeological bone. The biochemistry of inhibition is explored through chemical and enzymatic extraction methodology on archaeological material. We demonstrate a simple purification technique that provides a high yield of purified DNA (>95%) that can be used to address most types of inhibition commonly associated with the analysis of degraded archaeological and forensic samples. We present a new opportunity for the molecular analysis of archaeological samples preserved in the presence of metal ions, such as copper, which have previously yielded no DNA results.
Matheson
Carney D
CD
Department of Anthropology, Lakehead University, Thunder Bay, Canada. cmatheso@lakeheadu.ca
Marion
Travis E
TE
Hayter
Shana
S
Esau
Neal
N
Fratpietro
Renee
R
Vernon
Kim K
KK
D001842:Bone and Bones / Q000737:chemistry*; D002850:Chromatography, Gel / Q000379:methods*; D003300:Copper / Q000737:chemistry*; D004247:DNA / Q000737:chemistry / Q000302:isolation & purification*; D018732:Forensic Anthropology / Q000379:methods*; D005580:Fossils; D007477:Ions / Q000737:chemistry; D016133:Polymerase Chain Reaction; D017422:Sequence Analysis, DNA
D007477:Ions; D003300:Copper; D004247:DNA