Restriction site

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Restriction sites, or restriction recognition sites, are locations on a DNA molecule containing specific (4-8 base pairs in length[1]) sequences of nucleotides, which are recognized by restriction enzymes. These are generally palindromic sequences[2] (because restriction enzymes usually bind as homodimers), and a particular restriction enzyme may cut the sequence between two nucleotides within its recognition site, or somewhere nearby.

Function[edit]

For example, the common restriction enzyme EcoRI recognizes the palindromic sequence GAATTC and cuts between the G and the A on both the top and bottom strands, leaving an overhang (an end-portion of a DNA strand with no attached complement) known as a sticky end[2] on each end of AATT. This overhang can then be used to ligate in (see DNA ligase) 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.[2] 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.[3] 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.[4]

Applications[edit]

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

Databases[edit]

Several databases exist for restriction sites and enzymes, of which the largest noncommercial database is REBASE.[5][6]

See also[edit]

References[edit]

  1. ^ Russell, Peter J. (2006). iGenetics: A Mendelian Approach. Benjamin Cummings. ISBN 978-0805346664.
  2. ^ a b c Lehninger, Albert L.; Nelson, David L.; Cox, Michael M. (2008). Principles of Biochemistry (5th ed.). New York, NY: W.H. Freeman and Company. p. 305-306. ISBN 978-0-7167-7108-1.
  3. ^ Mousavi-Khattat, Mohammad; Rafati, Adele; Gill, Pooria (5 February 2015). "Fabrication of DNA nanotubes using origami-based nanostructures with sticky ends". Journal of Nanostructure in Chemistry. 5 (2): 177–183. doi:10.1007/s40097-015-0148-z.
  4. ^ Gao, Song; Zhang, Jiannan; Miao, Tianjin; Ma, Di; Su, Ying; An, Yingfeng; Zhang, Qingrui (28 March 2015). "A Simple and Convenient Sticky/Blunt-End Ligation Method for Fusion Gene Construction". Biochemical Genetics. 53 (1–3): 42–48. doi:10.1007/s10528-015-9669-x.
  5. ^ Roberts, Richard J.; Vincze, Tamas; Posfai, Janos; Macelis, Dana (2009-10-21). "REBASE—a database for DNA restriction and modification: enzymes, genes and genomes". Nucleic Acids Research. 38 (suppl_1): D234–D236. doi:10.1093/nar/gkp874. ISSN 0305-1048. PMC 2808884.
  6. ^ Roberts, Richard J.; Vincze, Tamas; Posfai, Janos; Macelis, Dana (2014-11-05). "REBASE—a database for DNA restriction and modification: enzymes, genes and genomes". Nucleic Acids Research. 43 (D1): D298–D299. doi:10.1093/nar/gku1046. ISSN 1362-4962. PMC 4383893.