Bacterial Transformation

Figure 1: Genetic modification example shown. Daniela Lin, StudySmarter Originals.

Figure 1 shows an example of what genetic modification can do. You might be wondering where we get the insect resistance gene copy from. Well, we get it from bacteria! We will elaborate on this more in-depth later in this article (under bacterial applications). Just understand that bacterial transformation can involve genetic modification on our part.

Although there are multiple techniques for genetic modification, they all generally involve deleting, adding, or turning off/on specific gene functions to give us desired traits in an organism.

Bacterial transformation steps

As explained above, transformation is the genetic change of a bacteria by the uptake of foreign DNA from the environment. Transformation can occur in nature but is rare and limited to certain bacteria.

On top of having chromosomal DNA, bacteria can also have plasmids. Plasmids are tiny, circular pieces of DNA that can be copied and transferred from bacteria to bacteria. Plasmids can also occur naturally in bacteria. Plasmids are the most common way scientists genetically modify bacteria as plasmids contain DNA sequences that can be edited in the lab.

Figure 2: How bacterial transformation can occur. Daniela Lin, StudySmarter Originals.

Figure 2 illustrates that when competent bacteria uptake foreign DNA from either their environment or other bacteria, the foreign DNA can either be 1) incorporated into the genome or chromosomal DNA or 2) incorporated into a plasmid. In nature, bacteria can transfer DNA using pili or a hairlike appendage; this process is called bacterial conjugation.

The typical steps involved in bacteria transformation in the lab are:

1. Desired colonies of bacteria are mixed with plasmids. These plasmids have been genetically modified to act as vectors.

• Vectors are things used to carry DNA segments into usually another bacterial cell (a host) as part of cloning or creating recombinant DNA.
• We modify a plasmid by cutting a plasmid open using restriction enzymes and putting the desired gene in. Once that's done, we need to rejoin the broken DNA sequence using DNA ligase.

2. We heat shock or electroporate bacteria to make them take up the plasmid. Heat shock or electroporation works because they both make the membrane more permeable to the plasmids by opening up the pores. (1)

3. We select bacteria that contain plasmids by putting them on antibiotic plates.

• Bacteria that contain plasmids expend more energy than bacteria that do not. This means that bacteria that do not have plasmids will grow faster than ones that do. This means we have to give the bacteria with plasmid advantages for them to want to keep their plasmid, and we do this by giving them all antibiotic-resistance genes.
• By placing all bacteria from steps 1 and 2 into antibiotic plates, we select only bacteria that contain plasmid because all the others die.
• Each bacterium with a plasmid multiplies into colonies.

4. We need to check colonies to identify which ones have the perfect plasmids. Once we do that, we can grow this colony and use it for protein or plasmid production.

• The reason we need to check for perfect plasmids is that sometimes when we create plasmids, we may get situations where the plasmid doesn't take up the desired gene, the gene goes in wrongly or backward, etc.

All steps 1-4 are illustrated in Figure 3 shown below for reference.

Figure 3: Transforming bacteria in the lab illustrated. Daniela Lin, StudySmarter Originals

Bacterial transformation diagram

So far, we've gone over bacterial transformation and how it relates to genetic modification. We've also looked at how bacteria transformation occurs 1) naturally and 2) in the lab. Now we can go over a diagram of how bacteria transformation was first discovered to understand the concept further.

Frederick Griffith first discovered the fact that bacteria could transform. He was the first to demonstrate that DNA could be horizontally transferred between organisms of the same generation instead of vertically (from parent to offspring). (2)

He performed the following experiments, which are shown in Figure 4:

1. He found that when he injected mice with a live strain S, they died since the strain was virulent (experiment 1).
2. He found they lived when he injected mice with a live strain R since the strain was not virulent (control group).
3. He found that the mice survived when he injected mice with a live strain R or a heat-killed strain S.
4. The problem was:

• The mice died when he injected them with a mix of live strain R and a heat-killed strain S. Curious, he isolated live bacteria from these dead mice and found that there was only strain S of bacteria! He injected this isolated S strain into live mice to confirm that they had died! This is shown in Figure 4 as experiments 2 and 3.

Figure 4: Griffith's transformation experiments illustrated. Daniela Lin, StudySmarter Originals.

Conclusion:

Griffith concluded that something had transferred from the heat-killed strain S to the live strain R. Today, this is known as Griffith's transformation principle or bacterial transformation.

Bacterial transformation purpose

The purpose of bacterial transformation in nature is to provide genetic diversity for bacterial cells allowing them to better adapt to a changing environment. Bacteria don't sexually reproduce, they asexually reproduce through binary fission.

Bacterial transformation applications

After understanding the purpose of transforming bacteria, we can now go over some of the applications in detail mentioned above.

DNA Cloning

• DNA Cloning is used to create many copies of a gene or DNA fragments that interest researchers.
• Scientists usually use DNA Cloning to synthesize recombinant versions of DNA to compare to normal genes in an organism to understand how they function initially. This is called genetic analysis.
• We can also use the transformed bacteria to make multiple plasmids and proteins to perform experiments in gene therapy. For instance, Synlogic, a biotech company, created "a designer bacteria" called SYNB1618 to attempt to cure PKU or Phenylketonuria, a rare disease that causes the build-up of phenylalanine. (3)

GMOs

• GMOs stand for genetically modified organisms, and we usually do it to foods to control pests, prevent crop loss, and provide longer shelf life, thereby driving down the cost.
• To produce a genetically modified organism, scientists need to identify a specific gene that produces the desired trait or function in an organism, such as insect resistance in figure 1.
• Once they have done this, you can isolate this DNA sequence and make many copies of it via DNA cloning in a bacteria (our host cell). Lastly, you transfer it to another organism or, in this example, another apple. If the protocol is done correctly, an insect-resistant fruit will be made.
• Like DNA cloning, GMOs also have strict regulations and checks to ensure that GMOs are toxic or harmful to our health.