Thermo Scientific T4 DNA Ligase catalyzes the formation of a phosphodiester bond between juxtaposed 5'-phosphate and 3'-hydroxyl termini in duplex DNA or RNA. The enzyme repairs single-strand nicks in duplex DNA, RNA, or DNA/RNA hybrids. It also joins DNA fragments with either cohesive or blunt termini, but has no activity on single-stranded nucleic acids.
T4 DNA Ligase requires ATP
as a cofactor.Highlights
• Active in Themo Scientific restriction enzyme, PCR, and RT buffers (when supplemented with ATP)
• Fast—sticky-end ligation is completed in 10 minutes at room temperature
• Supplied with PEG solution for efficient blunt-end ligationApplications
• Cloning of restriction enzyme generated DNA fragments
• Cloning of PCR products
• Joining of double-stranded oligonucleotide linkers or adaptors to DNA
• Site-directed mutagenesis
• Amplified fragment length polymorphism (AFLP)
• Ligase-mediated RNA detection (see
• Nick repair in duplex DNA, RNA or DNA/RNA hybrids
• Self-circularization of linear DNA.Includes
• T4 DNA Ligase
• 10X T4 DNA Ligase Buffer
• 50% PEG SolutionNotes
• Binding of T4 DNA Ligase to DNA may result in a band shift in agarose gels. To avoid this, incubate samples with 6X DNA Loading Dye & SDS Solution
at 70°C for 5 min or 65°C for 10 minutes and chill on ice prior to electrophoresis.
• The volume of the ligation reaction mixture should not exceed 10% of the competent cell volume in the transformation process.
• Prior to electro-transformation, remove T4 DNA Ligase from the ligation mixture using spin columns or chloroform extraction. The extracted DNA can be further precipitated with ethanol.
For Research Use Only. Not for use in diagnostic procedures.