Restriction Endonucleases

Restriction endonucleases (REs) cleave DNA at specific nucleotide recognition sequences. Restriction endonucle-ases are naturally occurring proteins found in and purified from bacteria. Each bacterial species contains one or more REs, each recognizing a unique sequence of base pairs in double-stranded DNA, called recognition sites (most commonly 4 to 8bp long). Bacteria use REs to digest and inactivate foreign DNA (such as bacteriophage DNA). The frequency of recognition sites in target DNA for any given RE is inversely proportional to the size of the recognition site. Some REs do not cleave DNA when their recognition sites are methylated; this can be useful in certain clinical laboratory applications such as detection of imprinted genes in genetic diseases or promoter hypermethylation in tumors. Some mutations occur at RE recognition sites and can be detected by a change in the RE digestion pattern of a PCR product or genomic DNA. Unique DNA restriction fragment patterns are generated by digestion with different REs, creating a range of DNA restriction fragment sizes. Fractionated using agarose gel electrophoresis. Restriction endonuclease digestion is commonly used as a component of clinical molecular tests.

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