Main menu

Long-read nanopore sequencing enables accurate confirmation of a recurrent PMS2 insertion–deletion variant located in a region of complex genomic architecture


Targeted next generation sequencing (NGS) is the predominant methodology for the molecular genetic diagnosis of inherited conditions. In many laboratories, NGS-identified variants are routinely validated using a different method, to minimize the risk of a false-positive diagnosis. This can be particularly important when pathogenic variants are located in complex genomic regions. In this situation, new long-read sequencing technologies have potential advantages over existing alternatives.

However, practical examples of their utility for diagnostic purposes remain scant. Here, we report the use of nanopore sequencing to validate a PMS2 mutation refractory to conventional methods. In a patient who presented with colorectal cancer and loss of PMS2 immunostaining, short-read NGS of Lynch syndrome-associated genes identified the recurrent PMS2 insertion-deletion variant, c.736_741delinsTGTGTGTGAAG (p.Pro246Cysfs*3). Confirmation of this variant using bidirectional Sanger sequencing was impeded by an upstream intron 6 poly(T) tract.

Using a locus-specific amplicon template, we undertook nanopore long-read sequencing in order to assess the diagnostic accuracy of this platform. Pairwise comparison between a curated benchmark allele (derived from short-read NGS and unidirectional Sanger sequencing) and the consensus nanopore dataset revealed 100% sequence identity. Our experience provides insight into the robustness and ease of deployment of “third-generation” sequencing for accurate characterisation of pathogenic variants.

Authors: Christopher M. Watson, Laura A. Crinnion, Jennifer Simmonds, Nick Camm, Julian Adlard, David T. Bonthron

Getting started

Buy a MinION starter pack Nanopore store Sequencing service providers Channel partners

Nanopore technology

Subscribe to Nanopore updates Resources and publications What is the Nanopore Community

About Oxford Nanopore

News Company timeline Sustainability Leadership team Media resources & contacts For investors For partners Working at Oxford Nanopore Current vacancies Commercial information BSI 27001 accreditationBSI 90001 accreditationBSI mark of trust
English flag