Circularization of Linear DNA

II. Circularization of Linear DNA

NOTE: Ensure that the DNA has been digested with DpnI previously. If it hasn’t, see step 6 in Site-Directed Mutagenesis.

A. Phosphorylation

1. Obtain a 0.6 mL tube and label it. Write CIRC below the DNA name add the reagents in the following order:

A. Phosphorylation

2. Incubate at 37C for 20 minutes . Write PHOS on the side of the tube.

1. In the same tube add the reagents in the following order:

B. Ligation

2. Incubate at room temperature for one hour. Write LIG on the side of the tube.

3. When done place the tube in the Freezer

Page updated

Report abuse