ISO 20397-3:2025

Overview - ISO 20397-3:2025 (Metagenomics, MPS)

ISO 20397-3:2025 is the first-edition International Standard that defines general requirements and guidance for metagenomics workflows using massively parallel sequencing (MPS). Intended for laboratories and research organizations, the standard covers end-to-end processes for metagenomics from primary sample handling through sequencing, bioinformatics analysis, and validation. It is Part 3 of the ISO 20397 series and complements Parts 1 and 2 on nucleic acid/library preparation and sequencing data quality.

Key SEO keywords: metagenomics standard, massively parallel sequencing (MPS), bioinformatics pipeline, library preparation, sample preparation.

Key Topics and Technical Requirements

The document specifies structured requirements across the complete metagenomics pipeline:

• Sampling strategy and sample handling
  • Primary sample types, stabilization, storage, transportation and extraction considerations
  • DNA/RNA isolation and nucleic acid sample quality assessment before library construction
• Nucleic acid library preparation
  • Library construction practices for DNA/cDNA/RNA suited to MPS platforms
• Design and sequencing strategy
  • Guidelines for short-read, long-read and hybrid sequencing approaches
  • Sample preparation, sequencing assessment and design review processes
• Database construction
  • Guidance for using public reference databases and building/curating self-built databases for taxonomic and functional annotation
• Bioinformatics analysis and reporting
  • Requirements for bioinformatics pipelines, sequence alignment, taxonomic identification lists, functional annotation, and report content
• Validation and verification
  • Pipeline and database validation using in silico controls and real sample controls; performance evaluation, verification and documentation
• Reporting and evaluation
  • Test report content and evaluation criteria for metagenomics outputs

Normative references include ISO 20397-1, ISO 20397-2, ISO/TS 24420 and ISO 20395.

Practical Applications and Users

ISO 20397-3 is useful for organizations performing metagenomics in:

• Environmental and ecological monitoring (microbial community profiling)
• Clinical microbiology and infectious disease research
• Agricultural and food-safety microbiome studies
• Public health surveillance and biodefense
• Core sequencing facilities and bioinformatics service providers

Primary users: laboratory managers, quality managers, molecular biologists, sequencing technicians, bioinformaticians, database curators and auditors.

Related Standards

• ISO 20397-1: Nucleic acid and library preparation
• ISO 20397-2: Quality evaluation of sequencing data
• ISO/TS 24420: Data processing for shotgun metagenomics
• ISO 20395: Performance evaluation of nucleic acid quantification methods

ISO 20397-3 provides a harmonized framework for robust, reproducible metagenomics workflows and for validating bioinformatics pipelines used in MPS-based microbial community analysis.