Home DNA Modifying Enzymes and Cloning Technologies 9°N™ DNA Ligase

9°N™ DNA Ligase
Gray property icon for NEBU recombinant 45 Gray property icon for heat inactivation no

9°N DNA Ligase has been reformulated with Recombinant Albumin (rAlbumin) beginning with Lot #10230152.

9°N DNA Ligase will ligate this substrate:

Nicked DNA/RNA



 

  • Can be used to ligate nicks in DNA while incubating at high temperatures
  • Extremely thermostable and can withstand PCR conditions
  • Will not ligate short 4 base overlaps (typical of restriction enzyme digests), but efficiently ligates 12 base pair overlaps
  • Isolated from a recombinant source
  • Not sure which ligase to choose?  Refer to our DNA and RNA Ligase Properties Chart or DNA Ligase Selection Chart

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DNA Ligase

DNA Ligation

Catalog # Cat # Concentration Conc Size List Price List Price Your Price Quantity
M0238S 40,000 units/ml 2,500 units $97.00 $97.00
Please enter a quantity for at least one size
  • 9° N™ DNA Ligase is a thermostable ligase that catalyzes the formation of a phosphodiester bond between the 5´-phosphate and the 3´-hydroxyl of two adjacent DNA strands that are hybridized and accurately paired, with no gap, to a complementary DNA strand.  9° N DNA Ligase uses ATP as a cofactor and it is active at elevated temperatures (45° C – 70° C).

    Product Source

    Purified from an E. coli strain containing the cloned ligase gene from the extremely thermophilic marine archaea Thermococcus sp.(strain 9°N). The archaea was isolated from a submarine thermal vent, at a depth of 2,500 meters, 9° north of the equator at the East Pacific Rise (1).
    This product is related to the following categories:
    DNA Ligases
    This product can be used in the following applications:
    Cloning Ligation

    • Reagents Supplied

      The following reagents are supplied with this product:

      NEB # Component Name Component # Stored at (°C) Amount Concentration
      • M0238S     -20    
          9°N™ DNA Ligase M0238SVIAL -20 1 x 0.063 ml 40,000 units/ml
          9°N DNA Ligase Reaction Buffer B0238SVIAL -20 1 x 1 ml 10 X

    • Properties & Usage

      Unit Definition

      (Cohesive End Unit) One unit is defined as the amount of enzyme required to give 50% ligation of the 12-base pair cohesive ends of 1 µg of BstEII-digested λ DNA in a total reaction volume of 50 µl in 15 minutes at 45°C. A cohesive end unit is equivalent to the nick-closing unit (1).

      Reaction Conditions

      1X 9°N DNA Ligase Reaction Buffer
      Incubate at 45°C

      1X 9°N DNA Ligase Reaction Buffer
      10 mM Tris-HCl
      600 µM ATP
      2.5 mM DTT
      2.5 mM MgCl2
      0.1% Triton® X-100
      (pH 7.5 @ 25°C)

      Usage Concentration

      40,000 units/ml

      Storage Buffer

      10 mM Tris-HCl
      50 mM KCl
      1 mM DTT
      0.1 mM EDTA
      200 µg/ml Recombinant Albumin
      50% Glycerol
      10 mM ammonium sulfate
      pH 7.4 @ 25°C

      Heat Inactivation

      No

      Unit Assay Conditions

      1X 9°N DNA Ligase Reaction Buffer and 20 µg/ml BstEII-digested λ DNA in a 50 µl reaction. After incubation at 45°C for 15 minutes, the reaction is terminated by addition of stop dye (50% glycerol, 50 mM EDTA and bromophenol blue), heated at 70°C for 10 minutes and then loaded on a 0.7% agarose gel. Due to the presence of ligase, the cos ends of BstEII-digested λ DNA will stay together after 70°C heat treatment.

    • Advantages and Features

      Application Features

      • Allele-specific gene detection using Ligase Detection Reaction and Ligase Chain Reaction (2,4).
      • Mutagenesis by incorporation of a phosphorylated oligonucleotide during PCR amplification (5).

    • Product Notes

      1. 9°N DNA Ligase is not a substitute for T4 DNA Ligase. The cohesive end unit is equivalent to the nick-closing unit of Barany et al.
      2. Incubate DNA and enzyme in 1X 9°N DNA Ligase Reaction Buffer at 45°C for 15 minutes or in a thermocycler with a program suited to the reaction described by Barany (1991) Genetic Disease Detection and DNA Amplification Using Cloned Thermostable Ligase. Proc. Natl. Acad. Sci. USA 88, 189-193. The reaction is stopped with a mixture of 50% glycerol, 50 mM EDTA, bromphenol blue.
      3. 9°N will not ligate short 4-base overlaps (typical of restriction enzyme digests), while it efficiently ligates 12-base pair overlaps.

    • References

      1. Thermococcus sp. (strain 9°N-7) isolated by Dr. Holger Jannasch (1991). Woods Hole Oceanographic Institute.
      2. Barany, F. (1991). Proc.Natl. Acad. Sci. USA. 88, 189-193.
      3. Takahashi, M. et al. (1984). J. Biol. Chem.. 259, 10041-10047.
      4. Barany, F. (1991). TheLigase Chain Reaction in a PCR World. (pp. 5-16, Cold Spring Harbor: Cold Spring Harbor Laboratory.
      5. Michael, S.F. (1994). Biotechniques. 16, 411-412.

  • Protocols, Manuals & Usage

  • Quality, Safety & Legal

    • Quality Control Assays

      Quality Control tests are performed on each new lot of NEB product to meet the specifications designated for it. Specifications and individual lot data from the tests that are performed for this particular product can be found and downloaded on the Product Specification Sheet, Certificate of Analysis, data card or product manual. Further information regarding NEB product quality can be found here.

    • Specifications & Change Notifications

      9°N DNA Ligase has been reformulated with Recombinant Albumin (rAlbumin) beginning with Lot #10230152. All subsequent (higher number) lots will contain rAlbumin.

      The Specification sheet is a document that includes the storage temperature, shelf life and the specifications designated for the product. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]

    • Certificate of Analysis

      The Certificate of Analysis (COA) is a signed document that includes the storage temperature, expiration date and quality controls for an individual lot. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]_[Lot Number]

    • Safety Data Sheets

      The following is a list of Safety Data Sheet (SDS) that apply to this product to help you use it safely.

    • Legal and Disclaimers

      Products and content are covered by one or more patents, trademarks and/or copyrights owned or controlled by New England Biolabs, Inc (NEB). The use of trademark symbols does not necessarily indicate that the name is trademarked in the country where it is being read; it indicates where the content was originally developed. The use of this product may require the buyer to obtain additional third-party intellectual property rights for certain applications. For more information, please email busdev@neb.com.

      This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.

      New England Biolabs (NEB) is committed to practicing ethical science – we believe it is our job as researchers to ask the important questions that when answered will help preserve our quality of life and the world that we live in. However, this research should always be done in safe and ethical manner. Learn more.

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