NextGenerationSequencing

Next Generation Sequencing (NGS) refers to high-throughput DNA and RNA sequencing technologies that perform parallel sequencing of millions to billions of fragments. Introduced in the mid-2000s, NGS dramatically increased speed and scale relative to Sanger sequencing, enabling large-scale genomic and transcriptomic analyses in research and clinical contexts.

NGS comprises multiple platforms. Illumina sequencing by synthesis dominates short reads, while earlier systems such as 454 pyrosequencing, SOLiD, and Ion Torrent helped establish parallel sequencing. More recently, third-generation approaches like PacBio single-molecule real-time sequencing and Oxford Nanopore Technologies provide long reads with limited amplification, aiding assembly and structural variant detection.

Typical workflows extract DNA or RNA, construct libraries, and generate millions of reads on a flow cell or nanopore device. Data are quality controlled, aligned to a reference, and analyzed for variants, gene expression, or methylation. Outputs include FASTQ, BAM/CRAM, and VCF files; metrics such as depth of coverage and read quality guide interpretation.

Applications span whole-genome sequencing, whole-exome sequencing, targeted panels, RNA sequencing, metagenomics, epigenomics, and single-cell analyses. NGS has transformed medical genetics and oncology, enabling diagnosis, pharmacogenomics, and precision medicine, while driving large-scale population genomics and evolutionary studies.

Limitations include data volume and computational requirements, platform-specific error profiles, and biases from library preparation. Costs per base have fallen, but data storage and analysis remain bottlenecks. Long-read technologies complement short reads by resolving repetitive regions and complex variants.