Gene Cloning for Testing Mutant Human Carbonic Anhydrase II Enzymatic Properties

Purpose

 We previously used PyMol modeling software to perform mutagenesis on specific amino acid residues within the active site of HCAII. In order to determine the changes in enzymatic properties of HCAII. We used several techniques including Gibson Assembly PCR to clone both the wildtype and mutant HCAII gene into pETblue-2 vector plasmid.

 We then transformed both the wildtype and mutant plasmids into E. coli. We used a heat shock procedure to transform the plasmids into the E. coli.