3.9 Replica Plating for Colony Screening

We will now examine how to select for transformed cells that harbor the plasmid of interest if that plasmid containes genes for antibiotic resistance. Think back to the plasmid pBR322. The original plasmid contains genes for resistance to amphicillin and tetracycline. Cells transformed with the original plasmid will be resistant to both of these antibiotics. If we cut the plasmid with BamH1, and insert our new gene at this site we will interrupt the gene for tetracycline resistance. If we transform cells with this new construct, the cells will not be resistant to tetracycline, but they will still be resistant to amphicillin.

Figure 21. Replica plating

Procedure:

Expected Result: The colonies of cells pressed onto normal medium will grow vigorusly as before and they represent the total number of cells that we plated.

In our example, cells plated on medium containing ampicillin will not grow unless they were transformed with either the original plasmid or the plasmid containing the inserted gene. The number of colonies tells us about the efficiency of transformation.