DNA Damage and PreCR
DNA is susceptible to many types of damage resulting from exposure to a variety of chemical or environmental reagents, manipulation or simply aging. The table below lists some possible DNA samples and the impact of the damage they may undergo.
Note: The extent of damage caused by exposure to different reagents can vary and its importance will depend on how the DNA is being used.
|Source of DNA||Potential Damage||Comments||References|
|Ancient DNA||abasic sites, deaminated cytosine, oxidized bases, fragmentation, nicks||Cytosine deamination has been reported to be the most prevalent cause of sequencing artifacts in ancient DNA.||Gilbert, M.T. et al. (2007) Nuc. Acid Res., 35, 1-10. PMID: 16920744
Hofreiter, M. et al. (2001) Nuc. Acid Res., 29, 4793. PMID: 11726688
|fragmentation, nicks||Nicks and fragmentation can increase the formation of artifactual chimeric genes during amplification.||Qiu, X. et al. (2001) Appl. Envir. Microbiol., 67, 880. PMID: 11157258|
|Source of Damage|
|abasic sites, oxidized bases, fragmentation, nicks||Ionizing radiation is used to sterilize samples.||Sutherland, B.M. et al. (2000) Biochemistry, 39, 8026. PMID: 10891084|
|Exposure to Heat||fragmentation, nicks, abasic sites, oxidized bases, deaminated cytosine, cyclopurine lesions||Heating DNA accelerates the hydrolytic and oxidative reactions in aqueous solutions.||Bruskov, V.I. (2002) Nuc. Acids Res., 30, 1354. PMID: 11884633|
|oxidized bases||Guanine is more sensitive to oxidation than the other bases and forms 8-oxo-G. 8-oxo-G can base pair with A making this damage potentially mutagenic.||Finnegan, M.T. (1995) Biochem. Soc. Trans., 23, 403S. PMID: 8566318|
|thymine dimers, (cyclobutane pyrimidine dimers) pyrimidine (6–4) pyrimidine photo products||UV trans-illumination to visualize DNA causes thymine dimer formation.||Cadet, J. et al. (2005) Mutat. Res., 571, 3-17. PMID: 15748634
Pfeifer, G.P. et al. (2005) Mutat. Res., 571, 19-31. PMID: 15748635
|fragmentation, nicks||Normal DNA manipulations such as pipetting or mixing can shear or nick DNA.|
|Dessication||fragmentation, nicks, oxidized bases||Mandrioli, M. et al. (2006) Entomol. Exp. App., 120, 239.|
|abasic sites, oxidized bases, deaminated cytosine, nicks, fragmentation||Long term storage in aqueous solution causes the accumulation of DNA damage.||Lindahl, T. et al. (1972) Biochemistry, 11, 3610 and 3618. PMID: 4626532|
|DNA-DNA crosslinks, DNA-protein crosslinks||Formaldehyde solution that has not been properly buffered becomes acidic, increasing abasic site formation.||Workshop on recovering DNA from formalin preserved biological samples. (2006) The National Academies Press.|
NEB now offers the PreCR Repair Mix (NEB #M0309), a blend of recombinant proteins designed to repair multiple types of damaged template DNA. It can be used in conjunction with any thermophilic polymerase.
The feature article, DNA Damage - the major cause of missing pieces from the DNA puzzle, provides a more in-depth look at the effects of different types of DNA damage.
|DNA Damage||Cause||Repaired by PreCR Repair Mix?|
|thymidine dimers||UV radiation||yes|
|Protein- DNA crosslinks||formaldehyde||no|