T4 DNA Ligase (EN11)

T4 DNA Ligase (EN11)

59,00 180,00 

ATP-dependent recombinant enzyme used for molecular cloning, site-directed mutagenesis, nick repair in duplex DNA, RNA or DNA/RNA hybrids, Ligation Mediated PCR; concentration 5 U/μl
EN11, EN11-050, EN11-250

SKU: EN11 Category:

Available Options:

Additional information

Unit

500 U, 2500U

Product Information

T4 DNA Ligase is an ATP-dependent recombinant enzyme isolated from Escherichia coli strain used to clone DNA. T4 DNA Ligase catalyzes the formation of a phosphodiester bond between juxtaposed 5′-phosphate and 3′-hydroxyl termini in duplex DNA or RNA. It will join both blunt-ended and cohesive-ended restriction fragments of DNA, as well as repair single-stranded nicks in duplex DNA, RNA or DNA/RNA hybrids.

Features:

  • ATP-dependent recombinant DNA ligases over-expressed in E. coli.
  • Available also in special formulation for very fast and efficient ligation of DNA fragments with compatible cohesive or blunt ends in 5 and 15 minutes respectively.

Applications:

  • Molecular cloning of PCR products or restriction fragments.
  • Joining double-stranded oligonucleotide linkers or adaptors to DNA.
  • Site-directed mutagenesis.
  • Nick repair in duplex DNA, RNA or DNA/RNA hybrids.
  • Self-circularization of linear DNA.
  • LM PCR methods (Ligation Mediated PCR).

Unit definition

One (Weiss) unit of T4 DNA Ligase catalyzes the conversion of 1 nmol of 32P from pyrophosphate into Norit-adsorbable material in 30 minutes at 37°C.
One Weiss unit is equivalent an approximately 200 cohesive end units.

Protocol & Manual

T4 DNA Ligase (EN11) - Manual

Technical Tips

  • The 10x Ligation Buffer and ATP Solution should be thawed and resuspended at room temperature.
  • For blunt-end ligations, use higher quantities of both the vector and the insert DNA.
  • For sticky (cohesive)-end ligations, we recommend heating both the vector and the insert DNA prior to ligation.
  • The electrotransformation efficiency may be improved by heat inactivation of the T4 DNA Ligase and purifi cation of the DNA by means of a spin column purification method (EXTRACTME DNA CLEAN-UP kit or
    EXTRACTME DNA GEL-OUT kit).
  • We recommend using a 3 – 10 molar excess of insert DNA over vector DNA. To calculate the optimal amounts of insert DNA in a ligation reaction,use the following equation:
    Example: If using 20 ng of a vector plasmid (4 kb), for a 5:1 molar ratio of insert : vector, you will require the following quantity of 1 kb insert:
  • The enzyme is inhibited by >200 mM NaCl or KCl concentrations.
  • Inactivate enzyme at 65°C for 10 minutes or at 70°C for 5 minutes.

FAQ

Quality & Safety

Quality control

The product was tested in ligation of HindIII-cut lambda DNA with a different amounts of enzyme. The product is free of unspecific DNA nucleases.

T4 DNA Ligase (EN11) - MSDS


T4 DNA Ligase (EN11) - CoA 1.2018 L.686771

Literature