NADseq

NADseq is a term used to describe sequencing-based approaches that investigate biological processes and molecules related to nicotinamide adenine dinucleotide (NAD+), a central cofactor in cellular metabolism. There is no single standardized protocol; various methods are grouped under the NADseq umbrella and often share the goal of profiling NAD+-associated features such as NAD-capped RNAs, NAD-binding proteins, or NAD+-dependent enzymatic activities.

Typical NADseq workflows involve selecting and enriching NAD-related targets from a biological sample, followed by library preparation and high-throughput sequencing. Enrichment strategies may rely on selective labeling, enzymatic tagging, or affinity capture to isolate NAD-associated species, after which the material is converted into sequencing-ready libraries. Bioinformatic analysis then aligns reads to reference genomes or transcriptomes, identifies NAD-enriched features, and interprets the data in the context of metabolic state and gene regulation.

Applications of NADseq include mapping the NAD-capped transcriptome to understand how NAD metabolism intersects with transcription, studying the dynamics of NAD+-dependent enzymes under metabolic or stress conditions, and exploring changes in NAD-related pathways across development or disease. The approach can provide insights into how cellular NAD+ levels influence RNA processing, protein modification, or signaling networks.

Limitations and considerations include potential biases from enrichment steps, the need for appropriate controls to distinguish genuine NAD associations from background, and the challenge of differentiating NAD-dependent signals from other nucleotide-related modifications. Related concepts include NAD-cap RNA profiling and broader NAD+-metabolite sequencing efforts.