Restriction and Ligation Enzymes

Overview[]

Restriction Enzymes and Ligation enzymes go hand in hand because in basic terms, one takes DNA strands apart and one puts DNA strands back together. These enzymes both have the ability to recognize and determine where to activate, typically at a desired location in a sequence. The coding of the ligation enzyme comes from the T4 Bacteriophage.

Restriction[]

Restriction enzymes recognize where on the sequence to cut and digest in order to create a single stranded “overhang.” This process is sometimes referred to as Gene Splicing. At this point, the fragment of DNA can either be replaced by another strand or even become the replacement strand at another location. This “overhang” is commonly termed the “sticky” or “cohesive” end because the unpaired bases (Adenine, Thymine, Cytosine, and Guanine) have the ability to bind via Ligation when they come in contact with a complimentary strand.

Ligation[]

The T4 DNA Ligase enzymes allow for the creation of covalent bonds between linear fragments of DNA. It is an ATP-dependent reaction, which means that the unpaired bases from the “sticky ends” of each fragment compliment each other and form Recombinant DNA. The 5’-phosphate end of the DNA strand covalently combines with the 3’-hydroxyl end of the DNA strand, thus completing the process.

Significance[]

The process of of Restriction and Ligation are very important tools in the field of genetics and genomics. It allows things such as recombinant DNA to be created and used for a plethora of applications. For instance, restriction and ligation can be used to insert a gene that codes for a particular protein of interest into a plasmid. This plasmid can be then inserted into the genomic DNA of bacteria and be transcribed by the organism to express the protein. This is an incredible breakthrough because the idea of inserting a gene that codes for a protein such as insulin could be revolutionary. Extracting this insulin from a bacterial source, then purifying it can lead to a whole new field of experimental pharmaceuticals.