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Products:Epigenetics and Nuclear Signaling >> DNA / RNA >> DNA Synthesis >> DNA Ligases
Overview
Product name
DNA Ligase IV protein (Active)
See all DNA Ligase IV products (2) ...
Protein description
Recombinant Full length DNA Ligase IV. Source: Escherichia Coli lambda lysogen NM 989.
Expression host
E. coli
Properties
Purification notes
Purified free of contaminating endonucleases and exonucleases. This product was tested in a mock cloning assay, which reveals any damage to the ligated DNA termini. Greater than 99.9% of the termini remain undamaged in this assay.
Biological activity
167,000 U/ml. Unit Defenition: One unit is defined as the amount of enzyme required to give 50% ligation of Hind III fragments of lambda DNA (5´ DNA termini concentration of 0.12 µM, 300µg/ml) in a total reaction volume of 20 µl in 30 minutes at 16°C in 1X T4 DNA Ligase Reaction Buffer: (50mM Tris-HCl (pH 7.8 at 25°C), 10mM Magnesium chloride, 10mM DTT, 1mM ATP, 25 µg/ml BSA and DNA (0.1 to 1 µM in 5´ termini)).
Form
Liquid
Storage instructions
Shipped at 4°C. Upon delivery aliquot and store at -20°C. Avoid freeze / thaw cycles.
Storage buffer
Preservative: None
Constituents: 50% Glycerol, 200µg/ml BSA, 50mM Potassium chloride, 10mM Tris HCl, 1mM DTT, 0.1mM EDTA, pH 7.4
Concentration
Concentration information loading...
Additional notes
T4 DNA Ligase can be inactivated by incubation at 65°C for 10 minutes.
Optimal ligation occurs at 16°C.
To dilute T4 DNA Ligase that will subsequently be stored at –20°C, 50% glycerol storage buffer should be used; to dilute for immediate use, 1x T4 DNA Ligase reaction buffer can be used.
T4 DNA Ligase is strongly inhibited by NaCl or KCl if the concentration is > 200mM.
Ligation of blunt-ended and single-base pair overhang fragments requires about 50 times as much enzyme to achieve the same extent of ligation as cohesive-end DNA fragments. Blunt-end ligation may be enhanced by addition of PEG 4000 (10% w/v final concentration) or hexamine chloride, or by reducing the ATP concentration to 50µM.
Research areas
Epigenetics and Nuclear Signaling >> DNA / RNA >> DNA Synthesis >> DNA Ligases
Applications
Show applications key
Our Abpromise guarantee covers the use of ab68649 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Functional Studies
Application notes
Biological Activity:
One Weiss unit is equivalent to circa 67 cohesive-end ligation units.
T4 DNA Ligase is strongly inhibited by NaCl or KCl if the concentration is > 200mM.
Ligation of blunt-ended and single-base pair overhang fragments requires about 50 times as much enzyme to achieve the same extent of ligation as cohesive-end DNA fragments. Blunt-end ligation may be enhanced by addition of PEG 4000 (10% w/v final concentration) or hexamine chloride, or by reducing the ATP concentration to 50µM.
Protein info
Function
Efficiently joins single-strand breaks in a double-stranded polydeoxynucleotide in an ATP-dependent reaction. Involved in DNA non-homologous end joining (NHEJ) required for double-strand break repair and V(D)J recombination. The LIG4-XRCC4 complex is responsible for the NHEJ ligation step, and XRCC4 enhances the joining activity of LIG4. Binding of the LIG4-XRCC4 complex to DNA ends is dependent on the assembly of the DNA-dependent protein kinase complex DNA-PK to these DNA ends.
Tissue specificity
Testis, thymus, prostate and heart.
Involvement in disease
Defects in LIG4 are the cause of LIG4 syndrome (LIG4S) [MIM:606593]. This disease is characterized by immunodeficiency and developmental and growth delay. Patients display unusual facial features, microcephaly, growth and/or developmental delay, pancytopenia, and various skin abnormalities.
Defects in LIG4 are a cause of severe combined immunodeficiency autosomal recessive T-cell-negative/B-cell-negative/NK-cell-positive with sensitivity to ionizing radiation (RSSCID) [MIM:602450]. SCID refers to a genetically and clinically heterogeneous group of rare congenital disorders characterized by impairment of both humoral and cell-mediated immunity, leukopenia, and low or absent antibody levels. Patients with SCID present in infancy with recurrent, persistent infections by opportunistic organisms. The common characteristic of all types of SCID is absence of T-cell-mediated cellular immunity due to a defect in T-cell development. Individuals affected by RS-SCID show defects in the DNA repair machinery necessary for coding joint formation and the completion of V(D)J recombination. A subset of cells from such patients show increased radiosensitivity.
Sequence similarities
Belongs to the ATP-dependent DNA ligase family.
Contains 2 BRCT domains.
Cellular localization
Nucleus.
Target information above from: UniProt accessionP49917
The UniProt Consortium
The Universal Protein Resource (UniProt) in 2010
Nucleic Acids Res. 38:D142-D148 (2010).
References for DNA Ligase IV protein (Active) (ab68649)
ab68649 has not yet been referenced specifically in any publications.
Publishing research using ab68649? Please let us know so that we can cite the reference in this datasheet
Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"