User:Tamra L. Fisher/Notebook/Research for Dr. Hartings/2012/08/14

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DNA ligation with T4 DNA Ligase

This will be done with the wildtype Asc Hb double-digested insert. The procedure is based off of the protocol on the NEB website. A NEBuffer and ATP will be used to see if maybe there is a problem with the T4 buffers we have.

  1. Mix 2μL of 10x NEBuffer 4, 5μL of the double-digested pQE-80-L-Kan vector, 10μL of the double-digested wt Asc Hb insert, 2μL 10mM ATP, and 1μL of T4 DNA Ligase.
  2. Place at 16°C overnight.
    • The thermocycler was used to maintain this temperature overnight.