Limitations of next generation sequencing
Reviewed by Ian Reekie

This is a report of a young female patient with clinically diagnosed retinitis pigmentosa who underwent genetic testing using an inherited retinal disease panel and whole exome sequencing by next generation sequencing (NGS) technology. This approach did not identify any pathogenic variants. Subsequently, the RP1 gene was sequenced by conventional Sanger sequencing, which identified a homozygous Alu insertion within the gene. The case report itself is of interest, but the principal reason for highlighting this report is the wider implications of the situation they discuss. Genetic medicine and its intersection with ophthalmology is increasingly important, in particular with regard to inherited retinal diseases. Increasing numbers of patients will be subjected to genetic testing, and the limits of the approaches taken must be understood by the clinicians seeing those patients. In this case, the inherent technical limitations of short read NGS meant that the relatively large Alu insertion was not identified at first. A working knowledge of common disease-causing variants in different populations is important for identifying where more in-depth investigation may be worthwhile. In this case it was known that an Alu element insertion into RP1 is a common disease-causing variant in Japanese patients, and conventional sequencing could be directly targeted to assess this gene.

Limitations of short-read NGS in detecting RP1 Alu insertions: a case emphasizing Sanger confirmation.
Hayashi T, Mizobuchi K, Higa N, et al. 
OPHTHALMIC GENETICS 
2026;47(2):204–8.