Two decades of metagenomic data on soil biodiversity in Germany frm samples preserved in the German Environmental Specimen Bank

The samples were collected as part of the routine soil sampling of the German Environmental Specimen Bank. The sampling sites include 11 locations in nine sampling regions of different ecosystem types (conurbation, agrarian, forestry and near-natural) in Germany. DNA was extracted with the ZymoBIOMICS™ DNA/RNA Miniprep Kit (Zymo Research, Freiburg, Germany). Libraries were prepared and purified according to the BEST protocol. Sequencing was performed on an Illumina NovaSeq 6000 PE150 platform at Novogene (Cambridge, UK), with a sequencing depth of 2 GB per sample. Sequences were automatically trimmed and quality checked with Autotrim v0.6.1 and FastQC. Taxonomic classification was performed sequentially using Kraken2 v2.1.2. For building the reference databases, representative or reference RefSeq assemblies were downloaded from NCBI (downloaded on January 13, 2023) for the following RefSeq categories: human, bacteria, and fungi and plants combined. For invertebrates, the newly created MetaInvert database was used, which contains genomes of 232 Central European invertebrate soil species (Collins et al. 2023). Each reference dataset was used to build a Kraken2 database with default k-mer size k = 35. For assignment, sequences were compared with the human database to control for potential contamination. Unassigned sequences were compared with the bacteria database, then to the fungi and plants database, and finally to the MetaInvert database. Results of the sequential assignments were merged. Raw sequencing data is available in NCBI as BioProject PRJNA1276286. More details here: https://www.biorxiv.org/content/10.1101/2025.07.21.665925v1