Cool E. Coli Transformation Lab

You think you've seen it all when it comes to bacteria? Well you ain't seen nothing yet!

TADAAA! Now you have! Feast your eyes on this--antibiotic resistant, glow-in-the-dark E. Coli. Sounds safe, right? Right.

In this lab we used the process of genetic transformation, which entails inserting a plasmid (insertable gene) into an organism's DNA to modify its traits. Here's how we did it...

We used four pre-prepared agar plates with the following contents...

1: pGLO-/LB (control)

2: pGLO-/LB/amp (control) 

3: pGLO+/LB/amp

4: pGLO+/LB/amp/ara

pGLO was the plasmid that we added to the organism's genetic material. pGLO+ indicates the presence of the plasmid and pGLO- indicates its absence. LB stands for Luria Broth, which prepared the E. Coli cells for the "log phase" of reproduction. Amp stands for ampicillin, an antibiotic that's a relative of penicillin. Only the cells that received the pGLO plasmid would be able to survive the wrath of the ampicillin. Ara stands for arabinose sugar, which provided the materials necessary for the pGLO+ cells to make their fluorescent protein.

First we started off with 2 small tubes, one contained plus Pglo another contained minus Pglo. Then we put 250 micro liters of transformation solution into each tube. We then picked up some colonies of ecoli and put them in each test tube. Then we put each of the test tubes on ice. We then added plasmid DNA to the +Pglo tube but not the -Pglo. After, we incubated them on ice for 10 minutes. Then we put them in heated water in order to break the plasma membranes so the plasmid DNA can enter the cells. We then added some nutrient broth to each tube so that the colonies could grow. Then we took the solution out from each tube and placed them in the four agar plates. The agar plate with -Pglo and lb showed that anything can live on the plate, the agar plate with -Pglo lb amp showed that the antibiotic works. The other two plates served the purpose of showing that the plasmid DNA was actually transferred to the host.

These are the four agar plates we used to grow the ecoli on

Incubation of the two test tubes on ice

Heat shocking to destroy the cell membranes of the bacteria

Here you ca see that the bacteria transformation was succesful because they are fluorescing.

Here are our controls, as you can see the plate containing the amp had nothing grow in it, which was what we were expecting.

Our lab was succesful in transforming the ecoli and at the end we also measured the transformation efficiencies of the bacteria for each of our lab groups.

As you can see we all got around the same number which indicates that we did the experiment successfully. Our transformation rate was 3.125 x 10^2 

In order to calculate this rate you have to go through the steps outline in the picutre below.