Today, we worked to insert the gene that makes jellyfish glow into the E. Coli bacteria. As this process involved many elements of using micropipettes, we first began with practicing our micro pipetting skills. We did this by performing various exercises, one of which involving creating micropipette art! In one of the exercises, we also used a micro centrifuge to centrifuge the colored liquids which we had pipetted on the sides of the tube together.
The first step in the gene injection process was to perform a ligation reaction with three tubes–two being controls and one meant for the bacteria. The next step was transformation in which we were required to create sterile environments at each of our stations, in which we were actually dealing with the bacteria. This process consisted of many intense and time dependent steps including thawing the bacteria on ice and putting the tubes in a hot water bath for a heat shock for exactly 30 seconds. This was one of the most critical steps in the process because if the bacteria sat in the hot water bath for one second too long, the bacteria would simply die because of the heat and the whole process would be defective as the bacteria would be dead. We then moved on to plating the solutions in the test tubes to be placed in the incubator, in which we will then check back and hopefully see that the bacteria is glowing indicating the process was successful.
After we got our plates out of the incubator, we gathered the data that is presented below. Our specific experiment was somewhat successful in that we performed a successful transformation and ligation reactions which only allowed 2 colonies of bacteria to be present in tube #3. When we took 50 uL out of the tube to be put in the Amp+ plate, we just happened to bring the only 2 colonies that were produced into the plate which would not make the bacteria glow because of an absence of Arabinose. Because of this, there was no bacteria left for the 50 uL to place in the plate containing both Amp+ and Arabinose. As a result, we were left with 2 colonies in the Amp+ plate and no colonies in the ++ plate containing both Amp+ and Arabinose–which did not glow. This likely occurred because of the small amount of bacteria present following the transformation and ligation reactions.
