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Home / Assay Kits

E. coli NAD+-dependent DNA ligase assay kit Plus

SKU : BHE16300073

This product includes 800 ul of 10 x buffer, 44 ul of 100x DNA substrate, 44 ul of 100 x NAD+, 21 ml of Reagent T, 10 ul of 500 x E. coli ligase and 220 ul of 10 x fluorescence dye for 100 assays.

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Mfr.No.	NLA100KE, NLA100K
Description	NAD+ -dependent DNA ligases are present in bacteria, some entomopox viruses and mimi virus. Since NAD+ -dependent DNA ligases are essential for bacterial growth, they are valuable targets for identifying novel antibacterial agents. The NAD+ -dependent DNA Ligase Assay Kit is to measure the DNA ligase product in which the diphosphodiester bond is formed at the single stand break of a duplex DNA substrate. The ligase reaction is monitored by the fluorescence intensity at 535 nm. The assay is in 96-well plate format and can be used for screening inhibitors of DNA ligases from gram-positive (such as S. pneumoniae) and gram-negative (such as E. coli) bacteria. The E. coli NAD+ -dependent DNA Ligase Assay Kit Plus (Catalog number NLA100KE) includes 800 µl of 10 x buffer, 44 µl of 100x DNA substrate, 44 µl of 100 x NAD+ (10 mM), 21 ml of Reagent T, 10 µl of 500 x E. coli ligase (500 nM) and 220 µl of 10 x fluorescence dye for 100 assays.
Drug target assays	Antibacterial drug targets

Product Manual	Datasheet
Protocols Description	Assay Protocol 1.Reagent preparation: 10 x DNA: dilute the 100 x DNA with water 100 x enzyme: 100 nM NAD+ -dependent DNA ligase diluted form the 500 nM stock with 1 x Buffer 1 mM NAD+ : dilute the 10 mM stock with water 1 x fluorescence dye: dilute the 10 x fluorescence dye 10-fold with water 2. Reaction: The total volume of each reaction mixture is 40µl including: 27.6µl of H2O, 4µl of 10 x buffer (Buffer LS), 4µl of 10 x DNA, 0.4µl of 100 x enzyme, and 4µl of 1 mM NAD+ . Incubate the reaction mixture at 37°C for 60 min. 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. 3. Detection: Add 200µl of Reagent T into the 40µl of reaction mixture. Then add 20µl of the 1 x fluorescence dye. Mix the reaction solution and incubate it for 15 min. Measure the fluorescence intensity at 535 nm using the excitation wavelength at 485 nm.

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Combination SKU	Supplier No.	Specifications	Your price	Quantity
BHE16300073-1	NLA100KE	E. coli ligase included	$613.14		Add to Cart
BHE16300073-2	NLA100K	enzyme not included	$460.35		Add to Cart

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