By Nicole Gadda
Fun rating: 5/5
Transformations get a full 5/5 fun rating from me! The possibilities are truly endless when it comes to transforming DNA into a cell. You can make any combination imaginable! Want your cells to fluoresce bright green? Want your cells to express a super cool protein you just discovered? You can simply transform them with DNA that’ll make that possible!
Difficulty rating: 3/5
I’d give transformations a 3/5 difficulty rating. The protocol itself is relatively easy to follow, but it can be long. In addition, any time you work with bacteria, contamination from other microbes is possible. Contamination can make your results difficult to interpret, so scientists must be super careful as to not contaminate their samples!
What is the general purpose?
A bacterial transformation is a technique used to insert foreign DNA into a bacterial host cell, with the goal of getting the host cell to express a specific gene or protein. For instance, if the inserted piece of DNA stores the information needed to build a green fluorescent protein (GFP), then your bacterial host cell will begin making GFP and glow green!
Why do we use it? Scientists can engineer our bacteria to express different genes by introducing foreign DNA sequences into the cell. We use transformations as a molecular biology tool to learn more about specific genes of interest.
How does it work?
Some bacterial cells naturally use transformation to uptake foreign DNA into their cell during harsh and extreme environments – separately from the hands of a scientist. This foreign DNA can come from other bacterial cells that died and lysed open, spilling their cellular contents out of their membrane upon dying. Bacterial cells use this process in order to adapt to their ever-changing environments. By incorporating new DNA into their genome, they give themselves the chance to acquire a new advantageous gene to help them survive in their environment.
In a lab setting, scientists can take advantage of this characteristic to design cells that have any DNA sequence of choice. The first step is to make their bacterial cells competent. A competent bacterial cell is one that is capable of taking up DNA from its environment. Even if a bacterial cell isn’t naturally competent, scientists can artificially make them capable of taking up DNA either through chemicals or electrical pulses.
To make cells competent through chemicals, cells are chilled in the presence of calcium phosphate. This makes the cell membrane permeable, which means small particles (like DNA) can pass through holes in the membrane and get inside the cell. Then, cells are subjected to a high temperature (42 degrees Celsius, or 107.6 degrees Fahrenheit) for less than a minute; this shocks the cells. Cells can also be made competent in a process called electroporation, where a machine sends electrical pulses, ultimately shocking the cells as well as making their membranes permeable.
Both high heat and electrical pulses result in high stress for the cell, mimicking a harsh and extreme environment where it would be advantageous for a bacterial cell in the wild to take up foreign DNA. When cells are subjected to high heat or electroporation, they’ll take up whatever foreign DNA you’ve added to their surrounding environment.
After this, the cells must rest in order to give them time to express the genes from the DNA uptaken by the cells. To ensure this foreign DNA stays in the chromosome, scientists will append an antibiotic resistance gene to it. By growing transformed cells on plates with an antibiotic, only colonies with resistance to that antibiotic (and therefore, with the segment of foreign DNA that confers resistance), will grow, ensuring the bacterial cells used in your experiments have the DNA segment you want!
