This tag could be histidine (His) tag, other marker peptides, or a fusion partners such as glutathione S-transferase or maltose-binding protein. To make this purification process easier, a purification tag may be added to the cloned gene. Low level of constitutive protein synthesis may occur even in expression vectors with tightly controlled promoters.Īfter the expression of the gene product, it may be necessary to purify the expressed protein however, separating the protein of interest from the great majority of proteins of the host cell can be a protracted process. protein is constantly expressed) in some expression vectors. Gene expression however may also be constitutive (i.e. The promoters used in expression vector are normally inducible, meaning that protein synthesis is only initiated when required by the introduction of an inducer such as IPTG. The promoter initiates the transcription and is therefore the point of control for the expression of the cloned gene. For example, prokaryotes expression vectors would have a Shine-Dalgarno sequence at its translation initiation site for the binding of ribosomes, while eukaryotes expression vectors would contain the Kozak consensus sequence. There are differences in the machinery for protein synthesis between prokaryotes and eukaryotes, therefore the expression vectors must have the elements for expression that are appropriate for the chosen host. These may include a promoter, the correct translation initiation sequence such as a ribosomal binding site and start codon, a termination codon, and a transcription termination sequence. coli, elements that allow them to be maintained in another organism, and these vectors are called shuttle vectors.įurther information: Transcription (genetics) and Translation (biology)Īn expression vector must have elements necessary for gene expression. Vectors used for protein production in organisms other than E.coli may have, in addition to a suitable origin of replication for its propagation in E. The cloning process is normally performed in Escherichia coli. The cloned gene may be transferred from a specialized cloning vector to an expression vector, although it is possible to clone directly into an expression vector. An example of the use of expression vector is the production of insulin, which is used for medical treatments of diabetes.Īn expression vector has features that any vector may have, such as an origin of replication, a selectable marker, and a suitable site for the insertion of a gene like the multiple cloning site. ![]() Escherichia coli is commonly used as the host for protein production, but other cell types may also be used. The expression of a protein may be tightly controlled, and the protein is only produced in significant quantity when necessary through the use of an inducer, in some systems however the protein may be expressed constitutively. The goal of a well-designed expression vector is the efficient production of protein, and this may be achieved by the production of significant amount of stable messenger RNA, which can then be translated into protein. ![]() The vector is engineered to contain regulatory sequences that act as enhancer and promoter regions and lead to efficient transcription of the gene carried on the expression vector. Expression vectors are the basic tools in biotechnology for the production of proteins. The vector is used to introduce a specific gene into a target cell, and can commandeer the cell's mechanism for protein synthesis to produce the protein encoded by the gene. A bacterial expression vector for expressing green fluorescent protein from the T7 promoter.Īn expression vector, otherwise known as an expression construct, is usually a plasmid or virus designed for gene expression in cells.
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