Library adaptors are short oligonucleotides that are used to prepare RNA and DNA samples for next-generation sequencing (NGS). Adaptors can also include additional functional elements, including sample indexes and unique molecular identifiers to improve the analysis of NGS libraries. Here, we describe control adaptors, termed CAPTORS, that include reference controls for NGS. CAPTORS can be seamlessly integrated during library preparation of RNA and DNA samples for the NGS workflow, and their encoded information retrieved during nanopore sequencing for analysis. We show how CAPTORS can measure sequencing accuracy of individual reads and pores during the sequencing experiment, evaluate the quantitative accuracy and complexity of metagenomic and RNA-seq libraries, and enable best-in-class normalisation between samples. These benefits can also be employed to improve clinical diagnosis, with CAPTORS able to model and correct systematic sequencing errors and improve the diagnosis of pathogenic BRCA1/2 variants in breast cancer. We propose that the integration of CAPTORS within the NGS workflow will improve the accuracy and reliability of output libraries, enable interoperable comparisons between samples and laboratories, and support the increasing translation of NGS for clinical diagnosis.
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