ISO/PWI 26863

Biotechnology —Requirements and guidelines for absolute quantification of RNA-seq using calibrators traced to SI

Scope

This document specifies the methodology for the absolute quantification of RNA-seq using spike-in reference materialswith metrological traceability to the International System of Units (SI).
This document is applicable to high-throughput RNA sequencing technologies. It provides a standardized framework for:
a)       The integration of reference materials into the transcriptomic sequencing workflow;
b)       The conversion of relative sequencing read counts into absolute transcript abundance (e.g., copy numbers per cell or per unit of total RNA);
c)        The quality control metrics required to ensure the metrological traceability and reliability of the quantification results.
This document is intended for use by research laboratories, clinical diagnostic facilities, biotechnology companies, and regulatory bodies involved in transcriptomic analysis, drug discovery, and precision medicine. It aims to facilitate the comparability of transcriptomic data across different laboratories, sequencing platforms, and experimental timeframes, thereby supporting the establishment of clinical thresholds and the advancement of quantitative biology.
NOTE          This document is specifically applicable to short-read sequencing technologies. While the principles of SI-anchored calibration are applicable to broader transcriptomic applications, long-read sequencing and other emerging technologies are currently excluded from the scope of this document. Future parts of this series may be developed to address these technologies as the methodology matures.