Removal of deaminated cytosines and detection of in vivo methylation in ancient DNA.
DNA sequences determined from ancient organisms have high error rates, primarily due to uracil bases created by cytosine deamination. We use synthetic oligonucleotides, as well as DNA extracted from mammoth and Neandertal remains, to show that treatment with uracil-DNA-glycosylase and endonuclease VIII removes uracil residues from ancient DNA and repairs most of the resulting abasic sites, leaving undamaged parts of the DNA fragments intact. Neandertal DNA sequences determined with this protocol have greatly increased accuracy. In addition, our results demonstrate that Neandertal DNA retains in vivo patterns of CpG methylation, potentially allowing future studies of gene inactivation and imprinting in ancient organisms.
Briggs
Adrian W
AW
Max-Planck-Institute for Evolutionary Anthropology, D-04103 Leipzig, Germany. briggs@eva.mpg.de
Stenzel
Udo
U
Meyer
Matthias
M
Krause
Johannes
J
Kircher
Martin
M
Pääbo
Svante
S
D000818:Animals; D018899:CpG Islands; D003596:Cytosine / Q000737:chemistry; D019175:DNA Methylation; D004260:DNA Repair; D003641:Deamination; D043224:Deoxyribonuclease (Pyrimidine Dimer); D015186:Hominidae / Q000235:genetics; D006801:Humans; D056665:Mammoths / Q000235:genetics; D009841:Oligonucleotides / Q000737:chemistry; D017422:Sequence Analysis, DNA / Q000379:methods*; D051981:Uracil-DNA Glycosidase
D009841:Oligonucleotides; D003596:Cytosine; D043224:Deoxyribonuclease (Pyrimidine Dimer); D051981:Uracil-DNA Glycosidase