Ligases are versatile and ubiquitous enzymes that join the 3’ hydroxyl and 5’ phosphate ends to form a phosphodiester bond, making them essential in nicked DNA repair, and ultimately genome fidelity. This biological role has also been extremely valuable in sealing the sticky ends of plasmids in molecular cloning. Their importance is attested by the fact most organisms have multiple ligases dedicated to specific pathways of repairing DNA. In eubacteria these ligases are powered by NAD+ rather than ATP. Each nick site requires 1 ATP or 1 NAD+ to power the ligase repair. order to join these fragments, the ligase progresses through three steps: