Template:SBB-Protocols Enz2

EcoRI/BamHI Digest of PCR Products

For PCR products, you will only digest a portion of your purified PCR product. Note that you must make a minor modification to the procedure if your DNA is shorter than 300bp (you add a little isopropanol to the ADB after melting).

• Set up the following reaction:

 8uL of eluted PCR product
 1uL of NEB Buffer 2
 0.5uL EcoRI
 0.5uL BamHI

• Incubate at 37 degrees on the thermocycler for 1hr
• Run an agarose gel, and melt with 600uL ADB buffer at 55 degrees. ****NOTE: If you are running short of time, this is an acceptable stopping point
• If the DNA is shorter than 300bp, add 250uL of isopropanol and mix prior to loading it on the column