The PreCR® Repair Mix is an enzyme cocktail formulated to repair damaged template DNA prior to its use in the polymerase chain reaction (PCR), microarrays or other DNA technologies. PreCR is active on a broad range of DNA damages, including those that block PCR (e.g. apurinic/apyrimidinic sites, thymine dimers, nicks and gaps) and those that are mutagenic (e.g. deaminated cytosine and 8-oxo-guanine). In addition, it will remove a variety of moieties from the 3´end of DNA leaving a hydroxyl group. The PreCR Repair Mix will not repair all damages that inhibit/interfere with PCR. For example, it will not repair 8-oxo-7,8-dihydro-2'deoxyadenosines or fragmented DNA. In fact, the ligase present in the mix is very active at sealing nicks in DNA but does not ligate blunt ends or nicks near a mismatch effectively. This limits the possibility of chimeric gene formation as a product of ligation. The PreCR Repair Mix can be used in conjunction with any thermophilic polymerase.
DNA is susceptible to many types of damage resulting from exposure to a variety of chemical or environmental agents, manipulation or simply aging. The DNA Damage and PreCR Table 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.
Table 1:
Types of DNA Damage
Figure 1: Repair of different types of DNA damage with the PreCR Repair Mix.
The gel shows trial amplifications from damaged DNA that was either not treated (-) or treated (+) with the PreCR Repair Mix. Type of DNA damage is shown. Note: heat treated DNA is incubated at 99°C for 3 minutes. Marker M is the 2-Log DNA Ladder (NEB #N3200 ).
This product is related to the following categories:
* PreCR Repair Mix components: Taq DNA Ligase, Endonuclease IV, Bst DNA Polymerase, Fpg, Uracil-DNA Glycosylase (UDG), T4 PDG (T4 Endonuclease V) and Endonuclease VIII.
Reaction Conditions: DNA template, 100 µM dNTPs, 1X ThermoPol Reaction Buffer, 1X NAD+ and 1 µl PreCR Repair Mix in a total reaction volume of 50 µl. Incubate at 37°C.
1X NAD+ Solution: 0.5 mM NAD+.
dNTPs are not included with the PreCR Repair Mix.
References
Diegoli TM, Farr M, Cromartie C, Coble MD, Bille TW. (2012). An optimized protocol for forensic application of the PreCR™ Repair Mix to multiplex STR amplification of UV-damaged DNA.. Forensic Sci Int Genet. Jul, 498-503. PubMedID: 22001155
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