molecular biology Molecular biology is a branch of biology that seeks to understand the molecule, molecular basis of biological activity in and between Cell (biology), cells, including biomolecule, biomolecular synthesis, modification, mechanisms, and interactio ...

, restriction sites, or restriction recognition sites, are regions of a

DNA Deoxyribonucleic acid (; DNA) is a polymer composed of two polynucleotide chains that coil around each other to form a double helix. The polymer carries genetic instructions for the development, functioning, growth and reproduction of al ...

molecule containing specific (4-8

base pair A base pair (bp) is a fundamental unit of double-stranded nucleic acids consisting of two nucleobases bound to each other by hydrogen bonds. They form the building blocks of the DNA double helix and contribute to the folded structure of both DNA ...

s in length) sequences of

nucleotide Nucleotides are Organic compound, organic molecules composed of a nitrogenous base, a pentose sugar and a phosphate. They serve as monomeric units of the nucleic acid polymers – deoxyribonucleic acid (DNA) and ribonucleic acid (RNA), both o ...

s; these are recognized by

restriction enzyme A restriction enzyme, restriction endonuclease, REase, ENase or'' restrictase '' is an enzyme that cleaves DNA into fragments at or near specific recognition sites within molecules known as restriction sites. Restriction enzymes are one class o ...

s, which cleave the DNA at or near the site. These are generally

palindromic sequence A palindromic sequence is a nucleic acid sequence in a double-stranded DNA or RNA molecule whereby reading in a certain direction (e.g. 5' to 3') on one strand is identical to the sequence in the same direction (e.g. 5' to 3') on the complemen ...

s (because restriction enzymes usually bind as

homodimer In biochemistry, a protein dimer is a macromolecular complex or protein multimer, multimer formed by two protein monomers, or single proteins, which are usually Non-covalent interaction, non-covalently bound. Many macromolecules, such as proteins ...

s), and a particular restriction enzyme may cut the sequence between two nucleotides within its recognition site, or somewhere nearby.

Function

For example, the common restriction enzyme

EcoRI EcoRI (pronounced "eco R one") is a restriction endonuclease enzyme isolated from species '' E. coli.'' It is a restriction enzyme that cleaves DNA double helices into fragments at specific sites, and is also a part of the restriction modificatio ...

recognizes the palindromic sequence GAATTC and cuts between the G and the A on both the top and bottom strands. This leaves an overhang (an end-portion of a

strand with no attached complement) known as a sticky end on each end of AATT. The overhang can then be used to ligate in (see

DNA ligase DNA ligase is a type of enzyme that facilitates the joining of DNA strands together by catalyzing the formation of a phosphodiester bond. It plays a role in repairing single-strand breaks in duplex DNA in living organisms, but some forms (such ...

) a piece of DNA with a complementary overhang (another EcoRI-cut piece, for example). Some restriction enzymes cut DNA at a restriction site in a manner which leaves no overhang, called a blunt end. Blunt ends are much less likely to be ligated by a DNA ligase because the blunt end doesn't have the overhanging base pair that the enzyme can recognize and match with a complementary pair. Sticky ends of DNA however are more likely to successfully bind with the help of a DNA ligase because of the exposed and unpaired nucleotides. For example, a sticky end trailing with AATTG is more likely to bind with a ligase than a blunt end where both the 5' and 3' DNA strands are paired. In the case of the example the AATTG would have a complementary pair of TTAAC which would reduce the functionality of the DNA ligase enzyme.

Applications

Restriction sites can be used for multiple applications in molecular biology such as identifying restriction fragment length polymorphisms (

RFLP In molecular biology, restriction fragment length polymorphism (RFLP) is a technique that exploits variations in homologous DNA sequences, known as polymorphisms, populations, or species or to pinpoint the locations of genes within a sequence. T ...

s). Restriction sites are also important consideration to be aware of when designing

plasmid A plasmid is a small, extrachromosomal DNA molecule within a cell that is physically separated from chromosomal DNA and can replicate independently. They are most commonly found as small circular, double-stranded DNA molecules in bacteria and ...

Databases

Several databases exist for restriction sites and enzymes, of which the largest noncommercial database is REBASE. Recently, it has been shown that statistically significant nullomers (i.e. short absent motifs which are highly expected to exist) in virus genomes are restriction sites indicating that viruses have probably got rid of these motifs to facilitate invasion of bacterial hosts.Nullomers Database
contains a comprehensive catalogue of minimal absent motifs many of which might potentially be not-yet-known restriction motifs.

References

{{DEFAULTSORT:Restriction Site Genetics techniques Molecular biology Restriction enzymes

Function

Applications

Databases

See also

References