Targeted next-generation sequencing panels for monogenetic disorders in clinical diagnostics: the opportunities and challenges

Abstract

Next-generation sequencing (NGS) will soon be used for clinically heterogeneous, inherited disorders and the increasing number of disease-causing genes reported. Diagnostic laboratories therefore need to decide which NGS methods they are going to invest in and how to implement them. We discuss here the challenges and opportunities of using targeted resequencing (TRS) panels for diagnosing monogenetic disorders. Of the different NGS approaches available, TRS panels offer the opportunity to sequence and analyze a limited set of predetermined target genes. At present, TRS panels offer better base-pair coverage, running times, costs and dataset handling than other NGS applications such as whole genome sequencing and whole exome sequencing. However, working with TRS panels also poses new challenges in variant interpretation, data handling and bioinformatic analyses. To optimize the analyses, TRS panel testing should be performed by bioinformaticians, clinicians and laboratory staff in close collaboration.

Keywords:

• coverage
• DNA diagnostics
• incidental finding
• next-generation sequencing
• sequence depth
• targeted resequencing panel
• variant interpretation

Acknowledgements

We thank Jackie Senior for editing the manuscript.

Financial & competing interests disclosure

The authors have no relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or pending, or royalties.

No writing assistance was utilized in the production of this manuscript.

Key issues

• In the coming years, next-generation sequencing will be widely introduced in DNA diagnostic laboratories and will replace traditional Sanger sequencing for the majority of diagnostic requests.

• Targeted resequencing panels (TRS) have better test characteristics with regard to base-pair coverage, running times, costs and handling of datasets than whole genome and whole exome sequencing approaches.

• TRS panels are relatively fast and efficient, so numerous disease-causing genes can be tested in multiple patient samples with short turn-around times.

• TRS diagnostic panels must be embedded in existing quality management systems that must also cover the handling and storage of genomic data and bioinformatics pipelines.

• Some of the challenges in using TRS diagnostic panels lie in detecting specific (trinucleotide) repeat sequences, large structural genomic rearrangements and deletions/duplications of one or more exons. Although software tools are being developed for these situations they have not yet replaced Sanger sequencing and multiplex ligation-dependent probe amplification.

• More challenges in the use of TRS diagnostic panels – as in the other two next-generation sequencing modalities – lie in interpreting and classifying variants. Limiting the number of genes tested to core disease gene lists will also limit the number of genetic variants observed.

• Developing and implementing TRS diagnostic panels should be done by laboratory staff and clinicians in close collaboration.