Complete gene cloning
What is DNA Cloning?
When you hear the term "cloning," you might think of the replication of whole organisms, such as the sheep Dolly. However, cloning something means creating an exact genetic copy of it. In molecular biology laboratories, what is most commonly cloned is a gene or a small fragment of DNA.
Cloning is the process of creating multiple identical copies of a specific piece of DNA. In DNA cloning, the gene or desired DNA fragment (for example, a gene related to an important human protein) is first inserted into a circular piece of DNA known as a plasmid. This is accomplished using enzymes that “cut and paste” DNA, resulting in a recombinant DNA molecule, or DNA composed of pieces from different sources.
A schematic representation of the transfer of a specific gene into a plasmid to create recombinant DNA.
Next, the recombinant plasmid is introduced into bacteria. The bacteria carrying the plasmid are selected and grown. During reproduction, they replicate the plasmid and pass it on to their offspring, producing copies of the DNA contained within it.
Applications of DNA Cloning
What are the benefits of creating multiple copies of a DNA sequence within a plasmid?
In some cases, many copies of DNA are needed to conduct experiments or to create new plasmids. In other instances, the DNA fragment encodes a useful protein, and bacteria are used as "factories" to produce the protein.
Through DNA cloning, you can express the gene for a protein in a host organism, allowing you to quickly produce large quantities of that protein. For example, insulin, hepatitis B vaccine, and many other proteins and vaccines have been produced this way, and this approach is currently being utilized for the development of COVID-19 vaccines and certain treatments for the virus. It can also be used to produce edible proteins.
Molecules of DNA created through cloning techniques are used for many purposes in molecular biology. Here is a short list of examples:
Production of Biopharmaceuticals: DNA cloning can be used to create human proteins with medical applications, such as the insulin mentioned above.
Gene Therapy: In some genetic disorders, patients lack a functional form of a specific gene. Gene therapy aims to provide the natural copy of the gene to the patient’s body cells.
Gene Analysis: In basic research laboratories, biologists often use DNA cloning to create synthetic and recombinant versions of genes, which helps them understand the natural genes of living organisms.
Steps to Use DNA Cloning for Protein Synthesis (e.g., Human Insulin) in Bacteria
Cut the Plasmid and Paste the Gene: This process involves restriction enzymes (which cut the DNA) and DNA ligase (which joins the DNA).
Introduce the Plasmid into Bacteria: Use antibiotic selection to identify the bacteria that carry the plasmid.
Cultivate a Large Quantity of Plasmid-Harboring Bacteria: Use them as "factories" to produce the protein. Harvest and purify the protein from the bacteria.