---+++ Dpn I digestion *Purpose* <br> To digest (Adeno) methylated GATC sites. Useful for removing cell-derived plasmid template from PCR samples. <br> *Use* <br> 1. Add 1µL of [[https://www.neb.com/products/r0176-dpni][DpnI]] to finished 50µL PCR reactions (or .5µL to 25µL reactions). Pipet or invert to mix. <br> 2. Incubate the mixture at 37°C for 1-2 hrs (depending on your paranoia or need to remove template DNA). Alternatively, the solution can be left overnight at room temperature. Periodic mixing may aid digestion (but is unnecessary). <br> 3. PCR cleanup or gel-purify the reaction for downstream processes. It's that easy! <br> *Other Notes* <br> Use in NEBuffer 4; however, it can be added directly to PCR sample.<br> Heat inactivate by incubating at 80°C for 20 minutes.<br> *Procurement* <br> Can be ordered directly from NEB. <br> Typically stored at -20°C; can be found in the common enzyme freezer box. <br> NEB unit definition: One unit is defined as the amount of enzyme required to digest 1 µg of pBR322 DNA (dam methylated) in 1 hour at 37°C in a total reaction volume of 50 µl. NEB buffer composition: 1X NEBuffer 4: <br> 20 mM Tris-acetate <br> 50 mM potassium acetate <br> 10 mM Magnesium Acetate <br> 1 mM Dithiothreitol <br> pH 7.9 @ 25°C <br>
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Topic revision: r3 - 2014-07-23 - SeanLeonard