Applications of Targeted Region Sequencing in Human Disease Studies and Clinical Care

Introduction to target region sequencing

Since the introduction of Sanger sequencing in 1977, genetic sequencing has been greatly improved with costs simultaneously falling. With the ability to rapidly produce large volumes of sequencing data, next-generation sequencing (NGS) enables researchers to obtain whole genome or targeted regions of samples. Targeted region sequencing is preferred by researchers and clinical doctors who focus on human diseases, which is based on the fact that genomics and epigenomics have identified a number of genes associated with disease. In a fast and economical manner, targeted region sequencing is able to obtain the complete catalog of disease genes, allowing us to see the differences among thousands of people to discover critical genes that cause human disease, such as cancer, heart disease, autism, and schizophrenia.

Targeted region sequencing combines both target enrichment and NGS technologies. You can view our article Current Techniques for Targeted Region Enrichment for a better understanding of target enrichment methods and recommended products. The available products on the market can meet the needs of diverse research. Alternatively, custom panels can be designed based on your ideas to specifically capture any genomic regions or genes of interest. In the following sequencing process, while whole genome sequencing only provides coverage of tens of fold, targeted region sequencing achieves coverage levels up to a 1,000-fold higher, enabling more confidential results. This article mainly discusses the use of target region sequencing in applications for human disease studies and clinical practice.

Application of targeted region sequencing in studying human disease

Most Mendelian disorders are caused by exonic or splice-site mutations that alter the amino acid sequence of associated genes. The causative mutations for genetic diseases are listed in the database of Mendelian disorders in humans ( Linkage analysis in family members is a traditional strategy of identification of disease-associated mutation. However, many diseases are rare, which poses challenges for linkage analysis in which data collections from large families are needed.

Whole exome sequencing (WES) has been proved as a robust and effective technique in replacement of linkage analysis. It can discover new disease-causing variants and verify benign single- and multiple-nucleotide polymorphisms in human populations. Since 85% of disease-related mutations are found in the protein coding regions, WES is especially productive to study human diseases. Other advantages of WES include: (i) reduced sample numbers; (ii) the capability of identifying causative mutations in diseases with genetic and phenotypic heterogeneity; (iii) cross checking for benign SNPs; (iv) the identification of breakpoints in balanced chromosome translocations and inversions. WES and other targeted region sequencing methods, followed by verification from nonconsanguineous families and functional and immunolabeling examinations, can discover critical causative genes from small pedigrees.

Application of targeted region sequencing in clinical sets

Most clinical applications of deep sequencing concentrate on known mutations to generate clear interpretable reports. Therefore, targeted sequencing using a panel of disease genes suit this aspect of clinical studies. But clinical applications of targeted region sequencing are faced with several limitations. First, there is no clear regulatory oversight. Second, data analysis and NGS errors remain to be solved. Third, there are controversial ethical issues. Even so, targeted region sequencing has been widely used for both research and clinical use.

  • Clinical specimens

The quality and amount of nucleic acid template are vital for the success of target enrichment and deep sequencing. The clinical sample types generally include fresh or frozen tissue that often yield high quality DNA and samples that often yield low quantity and/or compromised quality DNA like FFPE (formalin-fixed paraffin-embedded) tissue, decalcified FFPE tissue, bone marrow trephines, smears, and fine needle aspirates (FNA), plasma, and circulating tumor cells. Some studies have proved the usability of these clinical samples for NGS. And choosing the DNA isolation approach most-suited for the sample type is important to implement NGS testing. Several regulatory agencies such as Association for Molecular Pathology (AMP), American College of Medical Genetics (ACMG), College of American Pathologists (CAP), and US Centers for Disease Control and Prevention (CDC) have proposed guidelines for the validation, implementation, and performing NGS testing involved in clinical applications.

  • Tumor heterogeneity

Another issue needed to be considered is the clonal heterogeneity of tumors which generates the presence of multiple somatic mutations. Some methods such as the comparison and filtering of germline variants can provide better identification of the tumor heterogeneity. Additionally, similar challenges are presented by formalin-fixed artifacts. Do and Dobrovic (2015) once summarized the nature and origin of these artifacts and proposed approaches to overcome them.

If you are interested in our genomics services, please visit our website: for more information. We can provide a full package of genomics sequencing, including whole genome sequencing, whole exome sequencing, targeted region sequencing, mitochondrial DNA (mtDNA) sequencing, and complete plasmid DNA sequencing.


  1. Ballester L Y, Luthra R, Kanagal-Shamanna R, et al. Advances in clinical next-generation sequencing: target enrichment and sequencing technologies. Expert review of molecular diagnostics, 2016, 16(3): 357-372.
  2. Lin X, Tang W, Ahmad S, et al. Applications of targeted gene capture and next-generation sequencing technologies in studies of human deafness and other genetic disabilities. Hearing research, 2012, 288(1-2): 67-76.
  3. Ma R, Gong J, Jiang X. Novel applications of next-generation sequencing in breast cancer research. Genes & Diseases, 2017, 4(3): 149-153.
  4. Do H, Dobrovic A. Sequence artifacts in DNA from formalin-fixed tissues: causes and strategies for minimization. Clinical chemistry, 61(1), 64-71 (2015).
For Research Use Only. Not for use in diagnostic procedures.
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