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The concept of genes as carriers of phenotypic information was introduced in the early 19th century by Gregor Mendel, who later demonstrated the properties of genetic inheritance in peas. Over the next 100 years, many significant discoveries lead to the conclusions that genes encode proteins and reside on chromosomes, which are composed of DNA. These findings culminated in the central dogma of molecular biology, that proteins are translated from RNA, which is transcribed from DNA.
In 1952, Joshua Lederberg coined the term plasmid, in reference to any extrachromosomal heritable determinant. Plasmids are fragments of double-stranded DNA that can replicate independently of chromosomal DNA, and usually carry genes. Although they can be found in Bacteria, Archaea and Eukaryotes, they play the most significant biological role in bacteria where they can be passed from one bacterium to another by horizontal gene transfer, usually providing a context-dependent selective advantage, such as antibiotic resistance.
By the 1970s the combined discoveries of restriction enzymes, DNA ligase and gel electrophoresis together allowed for the ability to move specific fragments of DNA from one context to another, such as from a chromosome to a plasmid. This process of molecular cloning allowed scientists to break chromosomes down to study their genes individually, marking the birth of molecular genetics.
Today, scientists can easily study and manipulate genes and other genetic elements using specifically engineered plasmids, commonly referred to as vectors, which have become possibly the most ubiquitous tools in the molecular biologist’s toolbox.
Plasmids have become an essential tool in molecular biology for a variety of reasons, including that they are:
Plasmids used by scientists today come in many sizes and vary broadly in their functionality. In their simplest form, plasmids require a bacterial origin of replication, an antibiotic resistance gene, and at least one unique restriction enzyme recognition site. These elements allow for the propagation of the plasmid within bacteria, while allowing for selection against any bacteria not carrying the plasmid. Additionally, the restriction enzyme site(s) allow for the cloning of a fragment of DNA to be studied into the plasmid.
Below are some common plasmid elements:
|DNA sequence which allows initiation of replication within a plasmid by recruiting transcriptional machinery proteins|
|Allows for selection of plasmid-containing bacteria. Learn more in the Antibiotic reference.|
|Short segment of DNA which contains several restriction sites allowing for the easy insertion of DNA. In expression plasmids, the MCS is often downstream from a promoter.|
|Gene, promoter or other DNA fragment cloned into the MCS for further study.|
|Drives transcription of the target gene. Vital component for expression vectors: determines which cell types the gene is expressed in and amount of recombinant protein obtained.|
|The antibiotic resistance gene allows for selection in bacteria. However, many plasmids also have selectable markers for use in other cell types.|
|A short single-stranded DNA sequence used as an initiation point for PCR amplification or sequencing. Primers can be exploited for sequence verification of plasmids. See Addgene's sequencing primers.|
The combination of elements often determines the type of plasmid. Below are some common plasmid types:
If you are looking for an empty plasmid backbone for your experiment, see Addgene's empty backbone page for more information.