Figure 1

Protein synthesis as a function of magnesium glutamate and potassium glutamate in three different buffers CP, PEP and 3-PGA. (A) Luc synthesized as a function of magnesium glutamate with 5 nM plasmid pBEST-UTR1-Luc (CP, PEP and 3-PGA: 90 mM potassium glutamate, 1.5 mM of each amino acid, 2% PEG 8000). (B) Luc synthesized as a function of potassium glutamate with 5 nM plasmid pBEST-UTR1-Luc (CP: 2 mM magnesium glutamate, 1.5 mM of each amino acid, 2% PEG 8000; PEP: 1 mM magnesium glutamate, 1.5 mM of each amino acid, 2% PEG 8000; 3-PGA: 0 mM magnesium glutamate, 1.5 mM of each amino acid, 2% PEG 8000). (C) Kinetics of eGFP expression in the best conditions for the three different buffers with 5 nM plasmid pBEST-UTR1-eGFP (CP: 10 mM magnesium glutamate, 50 mM potassium glutamate, 1 mM of each amino acid, 2% PEG 8000; PEP: 9 mM magnesium glutamate, 30 mM potassium glutamate, 1 mM of each amino acid, 2% PEG 8000; 3-PGA: 6 mM magnesium glutamate, 40 mM potassium glutamate, 1 mM of each amino acid, 2% PEG 8000). Concentrations of magnesium glutamate and potassium glutamate reported here are the concentrations added to the cell-free reaction. The crude extract, dialyzed against the S30 buffer B, brings an additional 4.5 mM magnesium glutamate and 20 mM potassium glutamate to the cell-free reaction.