Expression vector
Encyclopedia
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
machinery ribosomal complexes
. The plasmid is frequently 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.
The goal of a well-designed expression vector is the production of large amounts of stable messenger RNA
, and therefore proteins. Expression vectors are basic tools for biotechnology
and the production of proteins such as insulin
that are important for medical treatments of specific diseases like diabetes.
After expression of the gene product, the purification of the protein is required; but since the vector is introduced to a host cell, the protein of interest should be purified from the proteins of the host cell. Therefore, to make the purification process easy, the cloned gene should have a tag. This tag could be histidine (His) tag
or any other marker peptide.
Expression vectors are used for molecular biology
techniques such as site-directed mutagenesis
. Cloning vectors, which are very similar to expression vectors, involve the same process of introducing a new gene into a plasmid, but the plasmid is then added into bacteria for replication purposes. In general, DNA vectors
that are used in many molecular-biology gene-cloning
experiments need not result in the expression of a protein.
Expression vectors must have expression signals such as a strong promoter, a strong termination codon, adjustment of the distance between the promoter and the cloned gene, and the insertion of a transcription termination sequence and a PTIS (portable translation initiation sequence).
s, especially plants used in agriculture
. Expression vectors have been used to introduce a vitamin A
precursor, beta-carotene
, into rice plants. This product is called golden rice
. This process has also been used to introduce a gene into plants that produces an insecticide
, called Bacillus thuringiensis toxin
or Bt toxin
which reduces the need for farmers to apply insecticides since it is produced by the modified organism. In addition expression vectors are used to extend the ripeness of tomatoes by altering the plant so that it produces less of the chemical that causes the tomatoes to rot. There has been controversy over using expression vectors to modify crops due to the fact that there are unknown health risks, possibilities of companies patenting certain genetically modified food
crops, and ethical concerns. Nevertheless, this technique is still being used and heavily researched. e.g. baculovirus is commonly used as expression vectors for insect cells.
Plasmid
In microbiology and genetics, a plasmid is a DNA molecule that is separate from, and can replicate independently of, the chromosomal DNA. They are double-stranded and, in many cases, circular...
that is used to introduce a specific gene
Gene
A gene is a molecular unit of heredity of a living organism. It is a name given to some stretches of DNA and RNA that code for a type of protein or for an RNA chain that has a function in the organism. Living beings depend on genes, as they specify all proteins and functional RNA chains...
into a target cell. Once the expression vector is inside the cell, the protein
Protein
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...
that is encoded
Genetic code
The genetic code is the set of rules by which information encoded in genetic material is translated into proteins by living cells....
by the gene is produced by the cellular-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...
and translation
Translation (genetics)
In molecular biology and genetics, translation is the third stage of protein biosynthesis . In translation, messenger RNA produced by transcription is decoded by the ribosome to produce a specific amino acid chain, or polypeptide, that will later fold into an active protein...
machinery ribosomal complexes
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....
. The plasmid is frequently engineered to contain regulatory sequences that act as enhancer
Enhancer (genetics)
In genetics, an enhancer is a short region of DNA that can be bound with proteins to enhance transcription levels of genes in a gene cluster...
and promoter regions and lead to efficient transcription of the gene carried on the expression vector.
The goal of a well-designed expression vector is the production of large amounts of stable messenger RNA
Messenger RNA
Messenger RNA is a molecule of RNA encoding a chemical "blueprint" for a protein product. mRNA is transcribed from a DNA template, and carries coding information to the sites of protein synthesis: the ribosomes. Here, the nucleic acid polymer is translated into a polymer of amino acids: a protein...
, and therefore proteins. Expression vectors are basic tools for biotechnology
Biotechnology
Biotechnology is a field of applied biology that involves the use of living organisms and bioprocesses in engineering, technology, medicine and other fields requiring bioproducts. Biotechnology also utilizes these products for manufacturing purpose...
and the production of proteins such as insulin
Insulin
Insulin is a hormone central to regulating carbohydrate and fat metabolism in the body. Insulin causes cells in the liver, muscle, and fat tissue to take up glucose from the blood, storing it as glycogen in the liver and muscle....
that are important for medical treatments of specific diseases like diabetes.
After expression of the gene product, the purification of the protein is required; but since the vector is introduced to a host cell, the protein of interest should be purified from the proteins of the host cell. Therefore, to make the purification process easy, the cloned gene should have a tag. This tag could be histidine (His) tag
Polyhistidine-tag
A polyhistidine-tag is an amino acid motif in proteins that consists of at least five histidine residues, often at the N- or C-terminus of the protein. It is also known as hexa histidine-tag, 6xHis-tag, and by the trademarked name His-tag...
or any other marker peptide.
Expression vectors are used for molecular biology
Molecular biology
Molecular biology is the branch of biology that deals with the molecular basis of biological activity. This field overlaps with other areas of biology and chemistry, particularly genetics and biochemistry...
techniques such as site-directed mutagenesis
Site-directed mutagenesis
Site-directed mutagenesis, also called site-specific mutagenesis or oligonucleotide-directed mutagenesis, is a molecular biology technique in which a mutation is created at a defined site in a DNA molecule. In general, this form of mutagenesis requires that the wild type gene sequence be known...
. Cloning vectors, which are very similar to expression vectors, involve the same process of introducing a new gene into a plasmid, but the plasmid is then added into bacteria for replication purposes. In general, DNA vectors
Vector (molecular biology)
In molecular biology, a vector is a DNA molecule used as a vehicle to transfer foreign genetic material into another cell. The four major types of vectors are plasmids, viruses, cosmids, and artificial chromosomes...
that are used in many molecular-biology gene-cloning
Cloning
Cloning in biology is the process of producing similar populations of genetically identical individuals that occurs in nature when organisms such as bacteria, insects or plants reproduce asexually. Cloning in biotechnology refers to processes used to create copies of DNA fragments , cells , or...
experiments need not result in the expression of a protein.
Expression vectors must have expression signals such as a strong promoter, a strong termination codon, adjustment of the distance between the promoter and the cloned gene, and the insertion of a transcription termination sequence and a PTIS (portable translation initiation sequence).
Some Common Uses
In recent years, expression vectors have been used to introduce specific genes in organismOrganism
In biology, an organism is any contiguous living system . In at least some form, all organisms are capable of response to stimuli, reproduction, growth and development, and maintenance of homoeostasis as a stable whole.An organism may either be unicellular or, as in the case of humans, comprise...
s, especially plants used in agriculture
Agriculture
Agriculture is the cultivation of animals, plants, fungi and other life forms for food, fiber, and other products used to sustain life. Agriculture was the key implement in the rise of sedentary human civilization, whereby farming of domesticated species created food surpluses that nurtured the...
. Expression vectors have been used to introduce a vitamin A
Vitamin A
Vitamin A is a vitamin that is needed by the retina of the eye in the form of a specific metabolite, the light-absorbing molecule retinal, that is necessary for both low-light and color vision...
precursor, beta-carotene
Beta-carotene
β-Carotene is a strongly-coloured red-orange pigment abundant in plants and fruits. It is an organic compound and chemically is classified as a hydrocarbon and specifically as a terpenoid , reflecting its derivation from isoprene units...
, into rice plants. This product is called golden rice
Golden rice
Golden rice is a variety of Oryza sativa rice produced through genetic engineering to biosynthesize beta-carotene, a precursor of pro-vitamin A in the edible parts of rice...
. This process has also been used to introduce a gene into plants that produces an insecticide
Insecticide
An insecticide is a pesticide used against insects. They include ovicides and larvicides used against the eggs and larvae of insects respectively. Insecticides are used in agriculture, medicine, industry and the household. The use of insecticides is believed to be one of the major factors behind...
, called Bacillus thuringiensis toxin
Bacillus thuringiensis
Bacillus thuringiensis is a Gram-positive, soil-dwelling bacterium, commonly used as a biological pesticide; alternatively, the Cry toxin may be extracted and used as a pesticide. B...
or Bt toxin
Bacillus thuringiensis
Bacillus thuringiensis is a Gram-positive, soil-dwelling bacterium, commonly used as a biological pesticide; alternatively, the Cry toxin may be extracted and used as a pesticide. B...
which reduces the need for farmers to apply insecticides since it is produced by the modified organism. In addition expression vectors are used to extend the ripeness of tomatoes by altering the plant so that it produces less of the chemical that causes the tomatoes to rot. There has been controversy over using expression vectors to modify crops due to the fact that there are unknown health risks, possibilities of companies patenting certain genetically modified food
Genetically modified food
Genetically modified foods are foods derived from genetically modified organisms . Genetically modified organisms have had specific changes introduced into their DNA by genetic engineering techniques...
crops, and ethical concerns. Nevertheless, this technique is still being used and heavily researched. e.g. baculovirus is commonly used as expression vectors for insect cells.