Documentation

O2A SEQUENCES ​

O2A SEQUENCES is the module of the O2A framework that manages metabarcoding samples, metadata, and sequences together: storing raw sequencing files (FASTQ), attaching harmonised sample metadata, running standardized analysis workflows, and generating submission-ready exports for public archives. Everything is organised by project: you select a project in the header, and files, workflows, and exports all scope to that project. Projects can be shared or made public, with role-based permissions. Sample metadata is held centrally in O2A SAMPLES (linked by shared persistent identifiers) and shared by every file linked to the same sample record.

O2A SEQUENCES holds active data only: once raw sequences are deposited in a long-term public archive they are removed locally, so the platform complements the public archives rather than duplicating them.

Getting started ​

  1. Create a project Create a new project, enter the project details, and click Save.

  2. Add members (optional) If others need access, add them as members and click Save. You can also do this later.

  3. Upload your files Go to Files and add your FASTQ files, either by dragging and dropping them or by selecting them from your file browser.

  4. Download the metadata table The file names are added to the metadata table automatically. Download the table - it already has one row per file.

  5. Fill in the table and upload it Fill in one row per file. Every file needs a row. Then upload the table.

  6. Changing metadata later Select the files and edit the metadata in the web interface, or upload a new metadata file to replace the old entries. Save

Notes ​

  • Upload the files first, then fill in the table. Otherwise the table has no file names in it.
  • Don't rename files after uploading. The metadata is matched by file name.

Metadata schema ​

The metadata schema conforms to the submission requirements of the European Nucleotide Archive (ENA) (www.ebi.ac.uk/ena), so that records created in O2A SEQUENCES can be submitted to ENA without further curation. It builds on the MIxS standard (Minimum Information about any Sequence) and the HARMONise schema (harmonise.awi.de), now incorporated into O2A SAMPLES, and every controlled field draws on the ENA/INSDC value lists. Fields designated mandatory by ENA are likewise required here; enforcing them is what keeps a record submission-ready for ENA and, via the GFBio broker, for the ASV Registry.

Required fields include: ​

  • sample_id — identifies the sample and links the file to its record, from O2A SAMPLES
  • sample_title -
  • field_event_id - from O2A SAMPLES
  • field_sample_description - Sample descripion, Free-text description of the sample: what it is, how and where it was taken. No fixed vocabulary. Example value: Seawater eDNA sample
  • field_sample_collection_date_begin - Sampling date in ISO 8601 format: YYYY-MM-DD (e.g. 2019-11-14).
  • taxon_id - Taxonomy ID according to NCBI
  • geogr_loc_lat - Geographic location latitude
  • geogr_loc_lon - Geographic location longitude
  • depth - sampling depth, how far down the sample was taken from the local surface (water / sediment / soil). Interval allowed. Example value: 10m or 10-20cm
  • elevation - sampling elevation
  • geog_loc_name - Country or sea name e.g. Arctic
  • geographic_location - Region and specific place name, i.e. a part of the country/sea. Example value: Fram Strait
  • env_broad_scale - Major environmental system (biome) as an ENVO term; several terms possible, separated by |. Example: oceanic epipelagic zone biome [ENVO:01000033]
  • env_medium - Environmental material immediately surrounding the sample, as an ENVO term. Example value: sea water [ENVO:00002149]
  • env_local_scale - Nearby entities, as an ENVO term; finer-grained than the broad scale. Example value: sea ice [ENVO:00002200]
  • subsample_name - Name of each fastq sample. Example, one core has been cut into several parts and each part has been sequenced. Then the core is sample and the parts subsamples, e.g. sample=seawater, subsample=30cm_depth, subsample=20cm_depth
  • sequencing_platform e.g. Illumina Myseq; please see ENA for all permitted platforms
  • library_strategy e.g. AMPLICON; please see ENA for all strategies
  • library_source e.g. metagenomic; please see ENA for all permitted sources
  • library_selection e.g. PCR; please see ENA for all permitted selection options
  • pcr-primer_fwd - primer sequence forward
  • pcr-primer_rev - primer sequence reverse

A record is considered complete only once all mandatory fields have been populated.

Import metadata ​

Metadata may be entered directly or imported from a prepared sheet.

Template download / upload. A metadata template (XLSX) can be downloaded from the Files workspace, completed offline, and re-uploaded.

Inline editing. Values may be edited directly in the metadata table: rows are selected to enable editing and then saved or discarded. A "set whole column" control applies a single value to all selected rows.

Because metadata reside in O2A SAMPLES and are keyed by sample ID, a single edit propagates to all files that share that sample.

Import sequencing data ​

Raw reads are uploaded as FASTQ files (.fastq, .fq, or gzip-compressed .gz). Files are added by dropping them onto the upload area or selecting them manually, and are staged within the active project. Forward and reverse reads (R1 / R2) are paired automatically into a single sample entry by their filename suffix; a sample lacking one member of the pair is flagged as unpaired.

The required per-sample metadata are supplied during upload, either individually or in bulk from a metadata sheet. Checksums (md5) are not provided in the template but are computed after upload. Each file records its size, md5 checksum, upload time, associated sample, read direction, and paired file.

Export data ​

Exports are accessed from the exports view of the active project. O2A SEQUENCES generates submission-ready exports for three destinations: GFBio, ENA, and the ASV Registry. Two export types are available: a tabular metadata / FASTQ export in GFBio format, and a calculated-ASV export derived from a completed workflow.

ENA ​

European Nucleotide Archive (ENA) (www.ebi.ac.uk/ena)

GFBio ​

GFBio (www.gfbio.org)

ASV Registry (workflow required) ​

The calculated-ASV export publishes the standardized, provenance-tracked ASV (Amplicon Sequence Variant) table produced by an analysis workflow to the ASV Registry. A workflow is eligible only once it has completed successfully and produced an ASV table; workflows that are still running, have failed, or lack an ASV table cannot be exported. Running a workflow is therefore a prerequisite for this export.

Workflow ​

A workflow is a single execution of a pipeline over a defined set of input files within a project. Each pipeline runs as a containerized Docker analysis; pipelines implemented with established workflow managers (e.g. Nextflow, Snakemake) run unmodified within these containers, and additional pipelines can be integrated through the same standardized interface.