T7 phage
Encyclopedia
Bacteriophage T7 is a bacteriophage
capable of infecting susceptible bacteria
l cells. It infects most strains of Escherichia coli
(including E. coli O157:H7
, a strain of E. coli which can cause foodborne illness
).
of the phage that is spherical with an inner diameter of 55 nm and a tail 19 nm in diameter and 28.5 nm long attached to the capsid. The head of the phage particle contains the roughly 40 kbp dsDNA genome of T7.
. T7 polymerase uses E. coli's endogenous thioredoxin
as a sliding clamp during phage DNA replication
(though thioredoxin normally has a different function). The sliding clamp functions to hold the polymerase onto the DNA, which increases the rate of synthesis; initiation, the process by which a polymerase binds to DNA, is time-consuming.
The T7 promoter sequence is used extensively in molecular biology
due to its extremely high affinity for RNA polymerase
and thus high level of expression.
. Chan et al. (2005) "refactored" the genome of T7, replacing approximately 12 kbp of its genome with engineered DNA. The engineered DNA was designed to be easier to work with in a number of ways: individual functional elements were separated by restriction endonuclease sites for simple modification, and overlapping protein coding domains were separated and, where necessary, modified by single base pair silent mutations.
Bacteriophage
A bacteriophage is any one of a number of viruses that infect bacteria. They do this by injecting genetic material, which they carry enclosed in an outer protein capsid...
capable of infecting susceptible 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 cells. It infects most strains of Escherichia coli
Escherichia coli
Escherichia coli is a Gram-negative, rod-shaped bacterium that is commonly found in the lower intestine of warm-blooded organisms . Most E. coli strains are harmless, but some serotypes can cause serious food poisoning in humans, and are occasionally responsible for product recalls...
(including E. coli O157:H7
Escherichia coli O157:H7
Escherichia coli O157:H7 is an enterohemorrhagic strain of the bacterium Escherichia coli and a cause of foodborne illness. Infection often leads to hemorrhagic diarrhea, and occasionally to kidney failure, especially in young children and elderly persons...
, a strain of E. coli which can cause foodborne illness
Foodborne illness
Foodborne illness is any illness resulting from the consumption of contaminated food, pathogenic bacteria, viruses, or parasites that contaminate food, as well as chemical or natural toxins such as poisonous mushrooms.-Causes:Foodborne illness usually arises from improper handling, preparation, or...
).
Virology
The virus has complex structural symmetry, with a capsidCapsid
A capsid is the protein shell of a virus. It consists of several oligomeric structural subunits made of protein called protomers. The observable 3-dimensional morphological subunits, which may or may not correspond to individual proteins, are called capsomeres. The capsid encloses the genetic...
of the phage that is spherical with an inner diameter of 55 nm and a tail 19 nm in diameter and 28.5 nm long attached to the capsid. The head of the phage particle contains the roughly 40 kbp dsDNA genome of T7.
Components
Gp5 (encoded by gene gp5) is T7 phage's DNA polymeraseDNA polymerase
A DNA polymerase is an enzyme that helps catalyze in the polymerization of deoxyribonucleotides into a DNA strand. DNA polymerases are best known for their feedback role in DNA replication, in which the polymerase "reads" an intact DNA strand as a template and uses it to synthesize the new strand....
. T7 polymerase uses E. coli's endogenous thioredoxin
Thioredoxin
Thioredoxin is a class of small redox proteins known to be present in all organisms. It plays a role in many important biological processes. In humans, it is encoded by the TXN gene. Loss-of-function mutation of either of the two human thioredoxin genes is lethal at the four-cell stage of the...
as a sliding clamp during phage DNA replication
DNA replication
DNA replication is a biological process that occurs in all living organisms and copies their DNA; it is the basis for biological inheritance. The process starts with one double-stranded DNA molecule and produces two identical copies of the molecule...
(though thioredoxin normally has a different function). The sliding clamp functions to hold the polymerase onto the DNA, which increases the rate of synthesis; initiation, the process by which a polymerase binds to DNA, is time-consuming.
The T7 promoter sequence is used extensively in 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...
due to its extremely high affinity for RNA polymerase
RNA polymerase
RNA polymerase is an enzyme that produces RNA. In cells, RNAP is needed for constructing RNA chains from DNA genes as templates, a process called transcription. RNA polymerase enzymes are essential to life and are found in all organisms and many viruses...
and thus high level of expression.
Applications in molecular biology
T7 has been used as a model in synthetic biologySynthetic biology
Synthetic biology is a new area of biological research that combines science and engineering. It encompasses a variety of different approaches, methodologies, and disciplines with a variety of definitions...
. Chan et al. (2005) "refactored" the genome of T7, replacing approximately 12 kbp of its genome with engineered DNA. The engineered DNA was designed to be easier to work with in a number of ways: individual functional elements were separated by restriction endonuclease sites for simple modification, and overlapping protein coding domains were separated and, where necessary, modified by single base pair silent mutations.
External links
- http://www.evergreen.edu/phage/docs/TOC_Forward.pdf
- http://au.expasy.org/viralzone/all_by_species/518.html ViralZone