Gene cloning is a DNA technology that enables scientists to produce multiple copies of the desired gene.
Often, scientists need to work with a gene in isolation. Naturally, genes are part of a very long DNA molecule with hundreds of other genes and, as in eukaryotes, even bigger non-coding regions. Isolating a gene from the effects of other interacting DNA regions allows understanding its unique function. Moreover, sometimes, the protein product of a particular gene is needed. All these necessitate gene cloning.
For gene cloning, a cloning vector is an essential requirement besides the gene (or any DNA region of interest) to be cloned. A cloning vector is a DNA molecule that can carry the gene of interest into the host organism and can multiply there. Bacterial plasmid DNA is used commonly as a cloning vector.
A plasmid (also called a vector) is a small circular DNA molecule that replicates independently of the chromosomal DNA. An essential feature of plasmid vectors is the ease with which a foreign DNA fragment can be introduced via the multiple cloning site (MCS). The MCS is a short DNA sequence containing multiple sites that can be cut with different commonly-available restriction endonucleases.
In cloning, the plasmid molecules can be used to provide a “folder” in which to insert the desired DNA fragment. Plasmids with foreign DNA inserted into them are called recombinant DNA molecules because they are created artificially and do not occur in nature. Plasmids are usually introduced into a bacterial host for proliferation. In the bacterial context, the fragment of DNA from the human genome (or the genome of another organism that is being studied) is referred to as foreign DNA (or a transgene) to differentiate it from the DNA of the bacterium, which is called the host DNA. As host bacteria proliferate, plasmids and the foreign DNA also multiply.
Proteins that are expressed from recombinant DNA molecules are called recombinant proteins. Not all recombinant plasmids are capable of expressing genes. The recombinant DNA may need to be moved into a different vector (or host) that is better designed for gene expression. Plasmids may also be engineered to express proteins only when stimulated by certain environmental factors so that scientists can control the expression of the recombinant proteins.
• Cloning small fragments of a genome allows specific genes, their protein products, and non-coding regions to be studied in isolation.
• A cloning vector is a DNA molecule that allows inserting a fragment of foreign DNA into them, carries this fragment into a host cell, and multiplies as host cells proliferate.
• A plasmid, which is used as a vector, is a small circular DNA molecule that replicates independently of the chromosomal DNA; it can be used to provide a “folder” in which to insert the desired DNA fragment.
• Recombinant DNA molecules are plasmids with foreign DNA inserted into them; they have created artificially as they do not occur in nature.
recombinant DNA: DNA that has been engineered by splicing together fragments of DNA from multiple species and introduced into the cells of a host.
plasmid: a circle of double-stranded DNA that is separate from the chromosomes, which is found in bacteria and protozoa.
restriction endonucleases: Commercially available enzymes that can target and cut a specific region of DNA; they are naturally found in bacteria
cloning vector: a DNA molecule that can carry the gene in a host, often plasmids in bacteria
eukaryote: an organism whose genetic material is organized into chromosomes in the cell nucleus
chromosomal DNA: DNA that is formed into a chromosome using proteins, often found in the nucleus
multiple cloning site: is a DNA region within a Plasmid that contains multiple unique Restriction enzyme cut sites
transgene: a gene which is artificially introduced into the genome of another organism.