Surveying the repair of ancient DNA from bones via high-throughput sequencing
A comparative analysis on DNA repair/recovery methodologies to increase the amount of recoverable material.
Sep 11, 2015
Authors: Nathalie Mouttham, Jennifer Klunk, Melanie Kuch, Ron Fourney, and Hendrik Poinar
BioTechniques, Vol. 59, No. 1, July 2015, pp. 19–25.
Abstract
DNA damage in the form of abasic sites, chemically altered nucleotides, and strand fragmentation is the foremost limitation in obtaining genetic information from many ancient samples. Upon cell death, DNA continues to endure various chemical attacks such as hydrolysis and oxidation, but repair pathways found in vivo no longer operate. By incubating degraded DNA with specific enzyme combinations adopted from these pathways, it is possible to reverse some of the post-mortem nucleic acid damage prior to downstream analyses such as library preparation, targeted enrichment, and high-throughput sequencing. Here, we evaluate the performance of two available repair protocols on previously characterized DNA extracts from four mammoths. Both methods use endonucleases and glycosylases along with a DNA polymerase-ligase combination. PreCR Repair Mix increases the number of molecules converted to sequencing libraries, leading to an increase in endogenous content and a decrease in cytosine-to-thymine transitions due to cytosine deamination. However, the effects of Nelson Repair Mix on repair of DNA damage remain inconclusive.