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| Assay target | |
| Assay area | |
| Target type | |
| Detection method | |
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| Main SKU |
Bacteria, some entomopox viruses and mimivirus carry NAD+-dependent DNA ligases. Bacteria need these ligases to grow, which makes them useful targets in the search for new antibacterial agents.
Kit vs Kit Plus
The only difference is the enzyme: NLA100K supplies all assay reagents without ligase, for use with your own NAD+-dependent DNA ligase, while NLA100KE adds 10 µl of 500 x E. coli ligase. Both are for 100 assays.
| Kit (enzyme not included), 100 assays (NLA100K) | 100 assays |
|---|---|
| Kit Plus (E. coli ligase included), 100 assays (NLA100KE) | 100 assays |
Kit Components
| Component | Kit (enzyme not included), 100 assays (NLA100K) | Kit Plus (E. coli ligase included), 100 assays (NLA100KE) |
|---|---|---|
| 10 x Buffer (Buffer LS) | 800 µl | 800 µl |
| 100 x DNA substrate | 44 µl | 44 µl |
| 100 x NAD+ (10 mM) | 44 µl | 44 µl |
| Reagent T | 21 ml | 21 ml |
| 10 x Fluorescence dye | 220 µl | 220 µl |
| 500 x E. coli ligase (500 nM) | Not included | 10 µl |
10 x Buffer LS: 200 mM HEPES pH 7.5, 400 mM NH4Cl, 20% glycerol, 20 mM DTT, 0.05% Tween-20, 5 mM EDTA, 0.6 mg/ml BSA, 80 mM MgCl2. Reagent T stops the reaction and denatures the DNA.
Assay Principle
The kit measures the ligation product, in which a phosphodiester bond is formed at a single-strand break of a duplex DNA substrate. After the reaction, Reagent T stops the reaction and denatures the DNA, and a fluorescence dye is added; ligase activity is read as fluorescence at 535 nm (excitation 485 nm).
Specifications
| Assay Target | NAD+-dependent DNA ligase activity |
|---|---|
| Target Type | Ligase |
| Assay Type | Enzyme Activity Assay; Inhibitor Screening Assay |
| Detection Method | Fluorescence |
| Detection Wavelength | Ex 485 nm / Em 535 nm |
| Plate Format | 96-well plate |
| Instrument Required | Fluorescence microplate reader |
| Assay Time | About 75 min (60 min reaction + 15 min detection) |
| Size | 100 assays |
| Storage | Please contact us to confirm storage conditions. |
| Shipping | Shipped frozen. |
Protocol
The manufacturer's protocol describes the procedure and reaction volumes.
- E. coli NAD+-dependent DNA ligase assay (PDF, see Documents): Covers catalog # NLA100K, NLA100KE.
Safety & Handling
Can’t find the exact format you need? We can help you source or customize this NAD+-dependent DNA ligase activity assay — including larger pack sizes, bulk reagents, bulk enzyme, or a protocol adapted to your samples. Click Talk to a Scientist, email support@biohippo.com, or explore our Research Services. Our team will be in touch shortly.
