T4 DNA Ligase catalyses the formation of a phosphodiester bond between the terminal 5′ phosphate and 3′ hydroxyl groups of duplex DNA or RNA. The enzyme efficiently joins blunt and cohesive ends and repairs single stranded nicks in duplex DNA, RNA or DNA/RNA hybrids.
T4 DNA Ligase Version Sizes Ligase Concentration Provided Buffer Low Concentration Ligase
1,500 U 7,500 U
2 U/µL 10X T4 DNA Ligase Buffer High Concentration Rapid Kit 1,500 U 7,500 U 10 U/µL 2X Rapid Ligation Buffer, 10X T4 DNA Ligase Buffer
10X T4 DNA Ligase Buffer is composed of 500 mM Tris-HCI, 100 mM MgCl 2, 50 mM dithiothreitol, 10 mM ATP, pH 7.6 @ 25 °C.
2X Rapid Ligation Buffer is composed of 132 mM Tris-HCI, 20 mM MgCl 2, 2 mM dithiothreitol, 2 mM ATP, 15% PEG, pH 7.6 @ 25 °C.
Figure 1: Purity The purity of the T4 DNA ligase is equal or greater than 95% as judged by SDS-polyacrylamide gel electrophoresis with Coomassie blue staining. Molecular weight markers are in Lane 1. T4 DNA ligase was loaded at 5 units (Lane 2) and 10 units (Lane 3).
Figure 2: Exonuclease and Endonuclease Activity Assay The T4 DNA ligase is free of detectable of exo- and endonuclease activities, as judged by agarose gel electrophoresis following incubation of 10 units of enzyme for 16 hours at 37 °C with 1 µg of HindIII-digested λ DNA (Lanes 3 and 4) and supercoiled pUC19 DNA (Lanes 5 and 6). Molecular weight markers are in Lanes 1 and 2.
The low concentration ligase includes NxGen T4 DNA Ligase at 2 U/µL and 10X T4 DNA ligase Buffer. The high concentration ligase includes NxGen T4 DNA Ligase at 10 U/µL and both 2X Rapid Ligation Buffer and 10X T4 DNA ligase buffer. Note - item ID 30243-3, NxGen T4 DNA Ligase (High Concentration) Kit, 10 U/µL, 40,000 units includes T4 DNA Ligase and 10X T4 DNA ligase buffer only.
