Figure 6

Introduction of a ribosome binding site improves production of the protein encoded by the downstream gene. (a) Four silent mutations at the 3’-end of cat were introduced to generate a more effective ribosomal binding site upstream of ppa. Sequences at the junction between cat and ppa are shown for the original (1) and modified (2) constructs. (b) Growth of cells on plates containing ampicillin (to force retention of the helper plasmid) and chloramphenicol (to confirm integration of the mutation cassette into the genome) after introduction of the mutation cassette by electroporation. The lack of colonies on the left indicates that construct 1 does not enable production of the essential protein Ppa. However, construct 2 allows growth, indicating that sufficient Ppa is produced.