Bacterial Transformation

A transformation is a technique used to incorporate a DNA fragment into bacteria so that the bacteria express that new trait. It can be done for a PCR colony test, for example, if the desired DNA fragment involves antibiotic resistance. The resulting bacteria which grow on the antibiotic agar plate will have this new, successful, DNA fragment because they were resistant to the antibiotic on the plate.

Transformations usually begin with a vial of bacteria from a specific strain, along with a 1μl micro centrifuge tube with the DNA sample of any concentration (ng/μl). Depending on the experiment or the experimenter, different amount of the DNA sample (ng) can be added. To calculate the exact amount of DNA solution to add to the bacteria in μl, you divide the desired amount of DNA by the concentration of the sample. The result is generally an extremely small number such as 0.023μl, in which case the p2 micropipette will be used which has a range of 0.01-1 μl. The DNA is added to the thawed bacteria and gently stirred with the pipette tip and let sit. After the bacteria sits, it is "heat shocked" in a 42 degree celcius water bath for 30 seconds. The heat will allow the bacteria to loosely open and allow the new DNA into them. This is then placed back onto ice.

After the DNA is incorporated into the bacteria, the bacteria is plated. Plating bacteria is my favorite thing to do in the lab. A volume of bacteria is added to an agar plate with appropriate antibiotic. This is done near a bunsen burner so that other bacteria do not take root (although they should not grow on the antibiotic anyways). Next, a glass pipette tip is sterilized with the flame, and bent in the middle and slightly at the end so that the resulting wand will not poke through the agar in any way. Using this, the bacteria is spread evenly around the agar plate, and the plate is put in the incubator to let the colonies grow.