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BglB mutant plasmids were incubated on ice, allowing the plasmid to permeate the E. coli's surroundings. Heat shock at 42 degrees C and immediate incubation on ice result in E. Coli's uptake of exogenous genetic material. The addition of terrific broth and incubation at 37 degrees C for an hour assist in the recovery of E. Coli and the incorporation of the plasmid. The recovered cells were spread evenly on agar plates and were incubated at 37 degrees C overnight. Multiple colonies on the agar plates should be visible within 12 hours.   A growth media for all the mutants, Growth media,  containing terrific broth and kanamycin solution, was created. A single colony was scraped from an agar plate and dipped into the growth media in a 15 mL falcon tube. The single colony contains the mutant plasmid, which contains a gene resistant to kanamycin. In the growth media, the transforming plasmid survives while the other bacteria are susceptible to this antibiotic. This process ensures that only the transformed cell is able to grow. The 15 mL falcon tubes were covered with tube seals and incubated with shaking for 37 degrees C.  [Purification]