Expression cloning
Encyclopedia
Expression cloning is a technique in DNA cloning that uses expression vector
s to generate a library of clones, with each clone expressing one protein. This expression library is then screened for the property of interest and clones of interest recovered for further analysis. An example would be using an expression library to isolate genes that could confer antibiotic resistance
.
in which the transcriptional and translational signals needed for the regulation of the gene of interest are included in the cloning vector. The transcriptional and translational signals may be synthetically created to make the expression of the gene of interest easier to regulate.
s. To this end, a bacteria
l expression clone may include a ribosome
binding site
(Shine-Dalgarno sequence
) to enhance translation of the gene of interest's mRNA, a transcription
termination sequence, or, in eukaryotes, specific sequences to promote the post-translational modification
of the protein product.
Expression vector
An expression vector, otherwise known as an expression construct, is generally a plasmid that is used to introduce a specific gene into a target cell. Once the expression vector is inside the cell, the protein that is encoded by the gene is produced by the cellular-transcription and translation...
s to generate a library of clones, with each clone expressing one protein. This expression library is then screened for the property of interest and clones of interest recovered for further analysis. An example would be using an expression library to isolate genes that could confer antibiotic resistance
Antibiotic resistance
Antibiotic resistance is a type of drug resistance where a microorganism is able to survive exposure to an antibiotic. While a spontaneous or induced genetic mutation in bacteria may confer resistance to antimicrobial drugs, genes that confer resistance can be transferred between bacteria in a...
.
Expression vectors
Expression vectors are a specialized type of cloning vectorCloning vector
A cloning vector is a small piece of DNA into which a foreign DNA fragment can be inserted. The insertion of the fragment into the cloning vector is carried out by treating the vehicle and the foreign DNA with a restriction enzyme that creates the same overhang, then ligating the fragments...
in which the transcriptional and translational signals needed for the regulation of the gene of interest are included in the cloning vector. The transcriptional and translational signals may be synthetically created to make the expression of the gene of interest easier to regulate.
Purpose
Usually the ultimate aim of expression cloning is to produce large quantities of specific proteinProtein
Proteins are biochemical compounds consisting of one or more polypeptides typically folded into a globular or fibrous form, facilitating a biological function. A polypeptide is a single linear polymer chain of amino acids bonded together by peptide bonds between the carboxyl and amino groups of...
s. To this end, a bacteria
Bacteria
Bacteria are a large domain of prokaryotic microorganisms. Typically a few micrometres in length, bacteria have a wide range of shapes, ranging from spheres to rods and spirals...
l expression clone may include a ribosome
Ribosome
A ribosome is a component of cells that assembles the twenty specific amino acid molecules to form the particular protein molecule determined by the nucleotide sequence of an RNA molecule....
binding site
Binding site
In biochemistry, a binding site is a region on a protein, DNA, or RNA to which specific other molecules and ions—in this context collectively called ligands—form a chemical bond...
(Shine-Dalgarno sequence
Sequence
In mathematics, a sequence is an ordered list of objects . Like a set, it contains members , and the number of terms is called the length of the sequence. Unlike a set, order matters, and exactly the same elements can appear multiple times at different positions in the sequence...
) to enhance translation of the gene of interest's mRNA, a transcription
Transcription (genetics)
Transcription is the process of creating a complementary RNA copy of a sequence of DNA. Both RNA and DNA are nucleic acids, which use base pairs of nucleotides as a complementary language that can be converted back and forth from DNA to RNA by the action of the correct enzymes...
termination sequence, or, in eukaryotes, specific sequences to promote the post-translational modification
Posttranslational modification
Posttranslational modification is the chemical modification of a protein after its translation. It is one of the later steps in protein biosynthesis, and thus gene expression, for many proteins....
of the protein product.
See also
- Molecular cell biology
- geneticsGeneticsGenetics , a discipline of biology, is the science of genes, heredity, and variation in living organisms....
- gene expressionGene expressionGene expression is the process by which information from a gene is used in the synthesis of a functional gene product. These products are often proteins, but in non-protein coding genes such as ribosomal RNA , transfer RNA or small nuclear RNA genes, the product is a functional RNA...
- Transcription (genetics)Transcription (genetics)Transcription is the process of creating a complementary RNA copy of a sequence of DNA. Both RNA and DNA are nucleic acids, which use base pairs of nucleotides as a complementary language that can be converted back and forth from DNA to RNA by the action of the correct enzymes...
- translationTranslationTranslation is the communication of the meaning of a source-language text by means of an equivalent target-language text. Whereas interpreting undoubtedly antedates writing, translation began only after the appearance of written literature; there exist partial translations of the Sumerian Epic of...
- λ phage
- pBR322PBR322pBR322 is a plasmid and was the first widely-used E. coli cloning vectors. Created in 1977, it was named after its Mexican creators, p standing for plasmid, and BR for Bolivar and Rodriguez....