T4 DNA Ligase MBG

• Catalyzes the formation of a phosphodiester bond between juxtaposed 5′-phosphate and 3′-hydroxyl termini
• Exhibits very fast and efficient ligation of DNA fragments with compatible cohesive or blunt ends

T4 DNA Ligase MBG 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 and repair single-stranded nicks in duplex DNA, RNA or DNA/RNA hybrids.

It is supplied with 10 mM Tris-HCl (pH 7.4), 0.1 mM EDTA, 50 mM KCl, 1 mM DTT, 50% (v/v) glycerol.

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 to approximately 200 cohesive end units.

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 and repair single-stranded nicks in duplex DNA, RNA or DNA/RNA hybrids.

• 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 before ligation.
• The electrotransformation efficiency may be improved by heat inactivation of the T4 DNA Ligase MBG and purification of the DNA through a spin column purification method.
• We recommend using a 3–10 molar excess of insert DNA over vector DNA.
• The enzyme is inhibited by >200 mM NaCl or KCl concentrations.
• Inactivate the enzyme at 65°C for 10 minutes or at 70°C for 5 minutes.

Quality Control

T4 DNA ligase activity is assayed in a reaction containing 1 µg of bacteriophage lambda DNA digested with HindIII, 1x T4 Ligation Buffer and varying amounts of enzyme for 20 minutes at 16°C. Results are assayed by agarose gel electrophoresis. The product is free of unspecific DNA nucleases.

Exonuclease and endonuclease activities were evaluated by gel electrophoresis following incubation of 1 µg of DNA with enzyme in a 50 µL volume for 4 hours at 37°C.

This is used for applications such as:

• Molecular cloning of PCR products or restriction fragments
• Site-directed mutagenesis
• Nick repair in duplex DNA, RNA or DNA/RNA hybrids
• Self-circularization of linear DNA
• LM PCR methods (Ligation Mediated PCR)

The acquisition of BLIRT by QIAGEN in the second quarter of 2022 paved the way for BLIRT’s expansion of R&D and manufacturing capabilities for customized and standardized recombinant enzymes and molecular biology reagents. Due to the transition, the rebranding process from BLIRT to QIAGEN is ongoing and the following resources still show BLIRT branding.
 
We assure you that the integration does not affect the quality, fit, form and function of the products.