metaSPAdes assembly pipeline

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Method synopsis

Standard metagenomics pipeline involving assembly with MEGAHIT, binning with multiple tools, dereplication, and taxonomic classification using GTDB-tk.

Synthesized from method descriptions across 4 papers using this protocol.

Procedure from a recent peer-reviewed implementation

Steps below were extracted from the most recent peer-reviewed implementation of this protocol in the corpus — Metagenome-assembled genomes for oligotrophic nitrifiers From a mountainous gravelbed floodplain (2025), Environmental Microbiology. Implementations in other papers (listed below) may differ.

Metagenome assembly and binning
Assemblies and co-assemblies were made using MEGAHIT (v1.3.3) and binned using contigs > 2000 nt and > 2500 nt, respectively, using MetaBAT2 (v2.12.1), MaxBin 2.0 (v2.2.6), and CONCOCT (v1.0.0) with multiple fastq files grouped by surface (50cm depth), middle depth (70-110 cm), and anoxic depths (130-150 cm). Bins were consolidated and filtered using metawrap bin_refinement to be above > 50% completeness and have < 10% contamination as calculated via CheckM (v1.1.3).
Quantities: Contigs > 2000 nt for assemblies, > 2500 nt for co-assemblies, >50% completeness and <10% contamination thresholdsDuration: Not specifiedConditions: Standard computational processing conditions
Equipment: MEGAHIT (v1.3.3), MetaBAT2 (v2.12.1), MaxBin 2.0 (v2.2.6), CONCOCT (v1.0.0), CheckM (v1.1.3)
MAG dereplication and taxonomic classification
All MAGs were dereplicated using dRep (v3.1.1) at 98% average nucleotide identity (ANI). Taxonomic classification for each representative MAG was made using the Genome Taxonomy Database toolkit (GTDB-tk) with the database release 07-RS207.
Quantities: 98% ANI threshold for dereplicationDuration: Not specifiedConditions: Standard computational conditions
Equipment: dRep (v3.1.1), GTDB-tk, GTDB database release 07-RS207