Editing Quick Change (section)

== Dpn I Digestion of the Amplification Products  ==

Note: It is important to insert the pipet tip below the mineral oil overlay when adding the Dpn I restriction enzyme to the reaction tubes during the digestion step or when transferring the 1 ÎŒl of Dpn I-treated DNA to the transformation reaction. Stratagene suggests using specialized aerosol-resistant pipet tips, which are small and pointed, to facilitate this process. 

1. Add 1 ÎŒl of the Dpn I restriction enzyme (10 U/ÎŒl) directly to each amplification reaction below the mineral oil overlay using a small, pointed pipet tip. 

2. Gently and thoroughly mix each reaction mixture by pipetting the solution up and down several times. Spin down the reaction mixtures in a microcentrifuge for 1 minute and immediately incubate each reaction at 37°C for 1 hour to digest the parental (i.e., the nonmutated) supercoiled dsDNA.