The human genome contains billions of DNA building blocks. While sequencing an entire genome provides a comprehensive view, it is often more practical and efficient to focus on specific DNA segments. This targeted approach allows researchers to delve deeply into relevant genetic information.
Understanding Target Enrichment NGS
Target enrichment next-generation sequencing (NGS) involves selectively isolating and sequencing specific regions of DNA from a larger sample. This method focuses the sequencing effort on areas of interest within a complex genome. It serves as a preparatory step before high-throughput sequencing, allowing for efficient and in-depth examination of chosen genetic areas. NGS technologies enable the simultaneous sequencing of millions of DNA molecules, making this focused analysis possible on a large scale. By enriching specific regions, scientists achieve higher sequencing depth, which is beneficial for detecting subtle genetic variations.
The Rationale for Focusing on Specific DNA Regions
Analyzing an entire genome, known as whole-genome sequencing (WGS), is costly due to the sheer volume of DNA. WGS also generates enormous data, much of which may not be relevant to a specific research question, complicating analysis. Achieving sufficient sequencing depth across the entire genome to reliably detect rare genetic variants can also be challenging. For instance, whole-exome sequencing, which targets all protein-coding regions, covers only about 1.5% of the human genome but captures most known disease-causing mutations.
Target enrichment NGS helps overcome these limitations. It significantly reduces sequencing costs by concentrating resources on smaller, more relevant portions of the genome. This targeted approach allows for much deeper sequencing coverage of specific genes or genomic areas, enhancing the ability to detect rare mutations or low-frequency variants. The resulting smaller datasets also simplify data analysis, leading to faster interpretation of genetic findings.
Key Approaches to Target Enrichment
Two main strategies are employed for target enrichment: hybridization-based capture and amplicon-based enrichment. Hybridization-based enrichment, also known as capture-based enrichment, uses specially designed short DNA or RNA sequences called “probes” or “baits.” These probes are complementary to the DNA regions of interest and bind to them through hybridization. Probes are often labeled, for example with biotin, allowing desired DNA fragments to be separated and “captured” using magnetic beads.
Amplicon-based enrichment relies on a process similar to polymerase chain reaction (PCR) to amplify target DNA regions. Specific short DNA sequences called “primers” are designed to bind to the ends of desired DNA segments. These primers then facilitate the creation of many copies of only the target regions, increasing their concentration in the sample. This method is faster and simpler than hybridization-based approaches, and performs well with low amounts of starting DNA.
Practical Applications in Research and Medicine
Target enrichment NGS has widespread utility in research and clinical medicine. In cancer research, it identifies specific mutations within tumor genes, aiding precise diagnosis, predicting disease progression, and guiding personalized therapies. For example, panels targeting hundreds of cancer-related genes provide comprehensive genomic profiles of tumors.
The technology also diagnoses genetic diseases by detecting known mutations or variants in genes linked to inherited conditions, such as cystic fibrosis or certain heart disorders. This focused sequencing provides high sensitivity for identifying disease-associated variants. In infectious disease surveillance, target enrichment NGS allows rapid sequencing of specific genes from pathogens like viruses or bacteria. This helps track outbreaks, identify drug resistance markers, and understand how these pathogens evolve over time. In pharmacogenomics, this method helps understand how an individual’s genetic makeup influences their response to medications, enabling tailored drug prescriptions.