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T4 DNA Ligase, 400 U/μL
Price
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The DNA ligase enzyme is extracted from bacteriophage T4. It can ligate either cohesive or blunt ends of DNA. It catalyzes
phosphodiester bond formation between 5' phosphate and 3' hydroxyl termini of duplex DNA or RNA. It requires Mg2+ and
ATP as co factors. It has molecular weight of ~68 kDa.
Source: An E. coli strain that carries gene which express T4 DNA ligase.
Quantity
Properties and Storage
- Unit Definition: One unit is defined as the amount of enzyme required to give 50% ligation of HindIII fragments of λ DNA (5 ́ DNA termini concentration of 0.12 μM, 300- μg/ml) in a total reaction volume of 20 μl in 30 minutes at 16 °C in 1X T4 DNA Ligation Buffer.
- Purity: 10% reducing SDS PAGE confirms ≥98% purity
- Protein Concentration: Concentration was measured using UV 280 nm
- Ligation assay: λ DNA (1 µg) digested with HindIII is incubated with 2000 U of T4 DNA ligase in a 10 µL reaction with 1× buffer at 37 °C for 16 hours. Agarose gel analysis shows efficiently ligated DNA with no degradation.
- RNase activity: A 10 µL reaction containing 40 ng of a 300-base ssRNA and ≥1 µL of T4 DNA ligase is incubated at 37 °C for 16 hours. More than 90% of the RNA remains intact, indicating no RNase contamination.
- DNase activity: A 50 µL reaction with λ-HindIII DNA and ≥2000 U of T4 DNA ligase incubated at 37 °C for 16 hours shows no detectable DNA degradation on agarose gel, confirming absence of DNase activity.
- Storage: Store at –20°C
- Spin the tubes briefly before use
Application
- Cloning of DNA fragments generated by restriction digestion
- Cloning of PCR product to desired vector
- Joining of adapters and linkers to DNA
Related Products
-
- Edna Rapid Ligation Kit
- Edna Restriction Enzymes
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