Bacterial Transformation

Bacteria are commonly used as host cells to make copies of DNA in the laboratory because they are easy to grow in large numbers. Your cellular machinery naturally carries out DNA replication and protein synthesis.

Amazing bacteria

Bacteria are incredibly versatile organisms that have the unique ability to take in foreign DNA and replicate (or copy) it. This gives them an evolutionary advantage and helps them survive changes in their environment. For example, bacteria can acquire DNA that makes them resistant to antibiotics. The bacterial genome is contained in a single circular chromosome. This genetic material floats freely in the cell, unlike in eukaryotic organisms where the genetic material is enclosed within a nuclear membrane. Bacteria can sometimes contain smaller circles of DNA, called plasmids, that have a much smaller number of genes. Plasmids can be exchanged between bacteria in a process called conjugation.

Use of plasmids in the laboratory

Plasmids can be used as vectors to transport foreign DNA into a cell. Once inside the cell, the plasmid is copied by the host cell’s own DNA replication machinery. In the laboratory, plasmids are specifically designed for bacteria to copy the DNA they contain.

Essential elements of the plasmid

Laboratory-engineered plasmids contain a small number of genes that aid transformation. These include:

  • An origin of replication. This is the specific sequence of nucleotides where DNA replication begins.
  • A multiple cloning site. This site contains recognition sites for specific restriction enzymes. These restriction enzymes can be used to “cut” the plasmid so that the foreign DNA can be “glued on” by ligation.
  • A resistance gene. This gene encodes a protein that bacteria need to survive in a particular growth medium, for example when a specific antibiotic is present.

The piece of DNA or gene of interest is cut from its original DNA source using a restriction enzyme and then glued into the plasmid by ligation. The plasmid containing the foreign DNA is now ready to be inserted into bacteria. This process is called transformation.

Bacterial transformation

Before the bacterial transformation, the bacteria are treated with a chemical called calcium chloride, which causes water to enter the cells and swell them. These swollen bacteria are known as competent bacteria. Plasmid DNA (containing the foreign DNA) is then mixed with the competent bacteria and the solution is heated. Plasmid DNA enters the bacterium through tiny pores created in the cell membranes. Once in the host cell, the plasmid DNA is copied many times by the bacteria’s own DNA replication machinery.

How to know if it worked?

After transformation, the bacteria are grown on a nutrient-rich food called agar. Only bacteria that contain a plasmid with antibiotic resistance will grow in the presence of antibiotics. For example, if bacteria are grown on agar containing the antibiotic ampicillin, only bacteria that have been transformed with a plasmid containing the ampicillin resistance gene will survive. The transformed bacteria can then be grown in large numbers. The DNA of interest, or the protein encoded by the DNA, can be isolated and purified.

When is transformation used?

Bacterial transformation is used:

  • To make multiple copies of DNA is called DNA cloning.
  • To make large amounts of specific human proteins, for example, human insulin, which can be used to treat people with type I diabetes.
  • Genetically modify a bacterium or other cell.