Restriction enzyme

Molecular cloning: A popular application of restriction enzymes has been in the generation of recombinant DNA molecules. The process involves the cutting of the donor DNA (usually a plasmid) and the vector DNA (usually a gene from another organism) by a restriction enzyme to yield compatible ends. These ends could be either ‘blunt’ or ‘sticky’. The two cleaved DNAs are joined together using an enzyme called DNA ligase to generate a recombinant DNA molecule. This recombinant DNA can, then, be introduced into a host organism for replication. For more details, refer Restriction Enzyme Cloning Manual

DNA mapping, also known as restriction mapping, involves the use of restriction endonucleases to obtain structural information of the DNA fragment or genome. Mapping involves determination of the order of the restriction enzyme sites in the genome. The DNA of interest, whose structure is to be determined, is cleaved with a series of restriction endonucleases to produce DNA fragments varying in size. These fragments are separated on an agarose gel to determine the structure of the DNA of interest.
Based on the known restriction enzyme sites of a specific DNA fragment, restriction endonucleases can be used to verify the identity of that DNA fragment.

Restriction landmark genomic scanning is a genome analysis method that utilizes a combination of restriction enzymes to visualize differences in methylation levels across the genome of a given organism. It is a useful technique to identify deviations from normal in any DNA. It is very effective in detecting hyper/hypomethylation in tumors, deletions or amplifications of genes, or changes in gene expression throughout the development of an organism.⁸

Gene sequencing: A large DNA molecule can be sequenced by digesting it with restriction enzymes and processing the resulting fragments through a DNA sequencer.

Restriction fragment length polymorphism (RFLP) involves the digestion of a DNA sample using restriction enzymes, separating these fragments based on length by gel electrophoresis and transferring them onto a membrane. These fragments are then bound to a radioactive or fluorescent labeled probe targeting specific sequences that are bracketed by restriction enzyme sites. An RFLP occurs when the resulting fragment lengths vary between individuals. Each individual has a unique pattern called the ‘biological bar code’. This technique was the first DNA profiling technique used in gene mapping, localization of genes for genetic disorders, determination of risk for disease, and paternity testing.⁹