Directed evolution of Ralstonia eutropha Soluble Hydrogenase enzyme

Target genes: HoxH and HoxY (hydrogenase genes)

Strategy:
Use site saturation mutagenesis at the active site of the SH, together with broad spectrum PCR error prone mutagenesis (?) to induce mutations in E-coli optimised synthetic DNA sequence. Then transfect Ralstonia eutropha culture with the DNA and screen for recombinant (endogenous genes replaced) hosts which exhibit improved hydrogenase activity (acid agar test).

1. Order materials (culture, primers for mutagenesis, genes)
2. Grow R eutropha and E coli cultures (->what pH does it like?)
3. Plate out in bicycle spoke fashion on acid agar plates with and without universal indicator to evaluate screening technique
4. Perform mutagenesis