Final Post!

After several weeks of work, I finally wrapped up my E. coli transformation experiment. The goal was to test how environmental stress, including high salt concentration, nutrient limitation, and low pH, would affect transformation efficiency using the pGREEN plasmid. This plasmid contains both a gene for ampicillin resistance and one for GFP, which causes the cells to glow under blue light if successfully expressed.

I prepared six experimental groups: a negative control (E. coli without plasmid on ampicillin plates), a positive control (E. coli with pGREEN on standard LB + ampicillin), three stress conditions (salt, nutrient-limited, and low pH), and a group plated on LB without antibiotic to confirm that the cells and plasmid were viable.

After incubation, I saw exactly what I expected on the control plates: the negative control showed no growth, and the non-antibiotic plates had full lawns of E. coli. What surprised me was that, at the 16-hour mark, the only groups showing visible colony formation were the stress groups, not the positive control. Each had small, pinhead-sized colonies beginning to form. They didn’t glow right away, but over time, I was able to see faint fluorescence under blue light, confirming that the plasmid had been taken up and expressed.

The lack of growth on the positive control plates suggests that I may have made a mistake in that group, possibly forgetting to add the plasmid or making a pipetting error. Still, seeing the stress groups succeed showed that transformation can occur even under challenging conditions, and that the cells were more resilient than I expected. In the end, despite a few setbacks, the experiment worked, and I was able to see glowing colonies, which was the most rewarding part of the entire process.