A simple and highly sensitive DNAzyme-based assay for nicotinamide adenine dinucleotide by ligase-mediated inhibition of strand displacement amplification

•The strategy carried out a visual assay for NAD+.•A strategy based on ligase-mediated inhibition on strand displacement amplification.•The colorimetric assay is simple, high sensitivity and high selectivity.•A novel platform for investigating cofactors, small molecules and DNA ligases.

Existing strategies for detecting nicotinamide adenine dinucleotide (NAD+) or other cofactors are commonly cumbersome and moderate sensitive. We report a novel DNAzyme-based visual assay strategy for NAD+ based on ligase-mediated inhibition of the strand displacement amplification (SDA). In the presence of NAD+, the SDA can be inhibited by the ligase reaction of two primers, which can initiate the SDA reaction in the case of no ligation, resulting in a dramatically decreasing yield of the SDA product, a G-quadruplex DNAzyme that can quantitatively catalyze the formation of a colored product. Therefore, the quantitative analysis for NAD+ can be achieved visually with high sensitivity. The developed strategy provides a simple colorimetric approach with high selectivity against most interferences and a detection limit as low as 50 pM. It also provides a universal platform for investigating cofactors or other related small molecules as well as quantifying the activity of DNA ligases.

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