Bacterial Tranformation

So behind. So, so, so very behind and I’m sorry.  But, that is beside the point.  This was a really cool lab.  It involved glowing and E. Coli.  Yes, E. Coli.  The gross little bacteria that is known for causing many of problems with fresh fruit and ground beef.  E. Coli is normally known for causing wicked diarrhea, nausea, vomiting, and fatigue.  But, that is actually just the stereotypical view of E. Coli.  Really, it is quite common.  As Mr. Ludwig notified us, there is E. Coli in your stomach right now, and are you sick?  No.  So, in this lab, we (still carefully) dealt with E. Coli.  How did we use it?  Well, we genetically engineered it by making it glow. (I’m not currently using a Mac so I cannot insert the cool guy emoji with little black sunglasses.)  Well, we made some of the bacteria glow.

Now, I know this doesn’t sound very easy, and in a way it wasn’t.  But, it was A LOT easier than you would think.  A certain kit that was provided to us granted us with everything needed and it’s instructions.  Here is that procedure accompanied with photos of my group performing this experiment.  Try to keep up here.

We first had to distinguish the active +pGLO tube and the inactive -pGLO tube.  Each tube required 250 ul of transformation solution and put both of those in these cool rack things and on ice.

Next, we had to take the E. Coli bacteria in the given holder cartage and extract a bit of it using this loops on sticks.  Each container of the pGLO stuff got an evenly spun dispense of the bacteria.  Place both on ice again.

Using a new sterile loop, we took a dip into the plasmid DNA in the solution.  You should be able to view a blowing-bubble-like film across the loop.  This film is inserted into the +pGLO tube only to make things different.  That tube is placed again on ice.

Next, was an incubation of the two tubes.  This was to still happen on ice, but on a larger scale than in a beaker.  During this step, we were to prep our agar plates.  They received the labels of LB-, LB/amp-, LB/amp+, and LB/amp/ara+.  The single ara plate contained food for the bacteria.

After the incubation was a rapid heat shock.  The tubes floated in a heat bath for 50 seconds, no more, and then were placed back onto the ice for two minutes.

Then, after all of that, all of the the LB nutrient broth was added to each tube.  Those sat for 10 minutes at room temperature.  Each tube was to be lightly tapped with our fingers after the sitting.

Each agar plate was then given a pipet hit of it’s specific tube substance.  Positive to positive, negative to negative.  The substances were spread around using a sterile loop for each.  Lastly, the amp plates were stacked on top of each other and placed in the actual incubator overnight.  The next we had to make our observations.

There was no bacteria growth on either of the negative plates.  But, there were white swirls on the LB- plate which might have just been from us.  But, the positive side was different.  The LB/amp+ had a good deal of bacteria, but, it did not glow.  With the LB/amp/ara+ had a slightly less amount of bacteria.  Though, it did glow.

In more simple terms, this is is how things turned out.

So, now that we have all of this information, what exactly does it mean?

With the LB- tube, you should expect no change.  This is because it was the control plate, or the plate in which no change was made.

What about the plates with the amp?  Normally, without the presence of an antibiotic (amp), you would expect the bacteria to not grow on the plate.  So, when the bacteria is modified genetically, it does not die as quickly as it normally would, causing new bacteria to grow more rapidly.

So then why does the LB/amp/ara+ bacteria glow?  Genetic engineering.  When adding the are sugar, it tells the DNA to go backwards and resist the antibiotic.  Though, also when it switches around, it turns on the same gene that makes the bacteria glow.

Overall, this was a pretty cool lab to experiment with.  We handled a strange bacteria and genetically engineered it to glow.  So, I think now you can trust my biology class to handle a career of genetic engineering and alter YOUR DNA so that your whole body can glow.  Orrrr, maybe not.