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Introduction

The term next-generation sequencing, or NGS, is the collective name for a diverse group of technologies with a common theme – ‘massively parallel sequencing’, where hundreds of thousands of fragments of DNA are sequenced in parallel.

Clinical applications of NGS

NGS is used clinically for whole genome sequencing (WGS), whole exome sequencing (WES), gene panel testing, and increasingly for single gene testing.

These genomic technologies have transformed the fields of oncology, rare disease, infectious disease and prenatal diagnostics, and are increasingly becoming an integral part of mainstream clinical practice across all healthcare specialties.

NGS is also extensively used for research, enabling an era of gene discovery and diagnosis of rare monogenic disorders, and the identification and diagnosis of genetic factors contributing to common complex disease.

NGS also underpinned the world-leading 100,000 Genomes Project, a pioneering study in which whole genome sequencing was first offered to NHS rare disease and cancer patients.

The form of NGS predominantly used by NHS clinical diagnostic labs at present is called short read sequencing. Increasingly, ‘long read sequencing’ technology is being explored for clinical applications, as this has several technical advantages over short read sequencing.

Advantages and limitations of NGS

Advantages

  • Multiple genes can be tested in a single assay, reducing the diagnostic odyssey associated with offering patients successive single gene tests.
  • Samples for different patients can be tested together in batches, improving cost effectiveness.
  • NGS has a high sensitivity for single nucleotide changes and small deletions or insertions (indels).
  • In some cases, copy number variation and even structural variants (WGS only) can also be detected.

Clinical situations in which NGS is particularly useful include those where many possible genes and variant types could be causing a patient’s features.

Limitations

  • Where a large number of genes are tested, careful filtering and interpretation of variants is required, as most are unlikely to be causal.
  • Incidental findings may be a concern where many genes are tested.
  • Large amounts of data are generated, requiring storage.
  • Some regions may not have enough sequencing reads. In any such ‘gaps in coverage’, variants might be missed.
  • Copy number variants may not be detected as reliably as single nucleotide variants/small indels.
  • Structural variants may be detected by WGS, but these would not usually be detected by targeted panel sequencing.
  • Currently, methylation changes are not detected, and repeat expansion disorders may be detected but are usually tested for separately.

For a summary table comparing the advantages and disadvantages of the different approaches to gene sequencing (gene panel/WES/WGS), view our Knowledge Hub article, Different approaches to gene sequencing.

Resources

For clinicians

References:

Tagged: Sequencing, Technologies

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  • Last reviewed: 10/06/2022
  • Next review due: 10/06/2024
  • Authors: Dr Amy Frost, Dr Julia van Campen
  • Reviewers: Professor Barbara Jennings, Dr Siobhan Simpson, Professor Kate Tatton-Brown