NGS

Next-Generation Sequencing (NGS) refers to a set of high-throughput DNA sequencing technologies that generate millions to billions of sequences in parallel, enabling rapid analysis of entire genomes, exomes, transcriptomes, and more. NGS platforms typically produce short reads, although some long-read technologies can yield tens of kilobases per read. The approach dramatically increases sequencing capacity and reduces cost per base compared with traditional Sanger sequencing.

Common NGS platforms include Illumina’s sequencing by synthesis systems, Ion Torrent sequencing, and SOLiD, which primarily generate short reads. Long-read platforms such as Pacific Biosciences’ Single Molecule Real-Time (SMRT) sequencing and Oxford Nanopore Technologies provide longer reads with different error profiles. Underlying methods vary, but most rely on library preparation to create DNA fragments or cDNA, followed by amplification or real-time observation of nucleotide incorporation, and large-scale parallel sequencing.

The typical workflow involves sample preparation and library construction, sequencing to produce raw reads, and bioinformatic analysis. Data are stored in formats such as FASTQ for reads, BAM/CRAM for alignments, and VCF for variants. Common applications include whole-genome sequencing, whole-exome sequencing, targeted gene panels, RNA sequencing, metagenomics, and epigenetic studies using bisulfite sequencing or chromatin accessibility assays.

NGS has transformed research and medicine by enabling rapid discovery, personalized genomics, and large-scale population studies. Its advantages include high throughput and decreasing costs, while limitations include shorter read lengths for many platforms, platform-specific error profiles, computational demands, and data management challenges. Ethical, legal, and regulatory considerations are also important in clinical contexts.