Soil Microbiome
Metagenomics shotgun sequencing was performed by EzBiome (Gaithersburg, MD,USA). Concentration of genomic DNA was measured using the Qubit FluorometerdsDNA DNA quantification System (ThermoFisher, USA). 150ng of genomic DNA was used for library construction using NEBNext Ultra II FS DNA Library Prep Kit for Illumina. Briefly, gDNA was enzymatically sheared, DNA fragment ends wererepaired, 3 adenylated, and ligated to adapters according to the manufacturers instructions. The resulting adapter-ligated libraries were PCR-amplified using the following protocol: an initial denaturation step performed at 98C for 45s followed by5 cycles of denaturation (98C, 15 s), annealing (60C, 30 s) and extension (72C, 30sec), and a final elongation of 1 min at 72C. PCR product was cleaned up from thereaction mix with Mag-Bind RxnPure Plus magnetic beads (Omega Bio-tek, Norcross,GA).The libraries were quantified and qualified using the D1000 ScreenTape on anAgilent 2200 TapeStation instrument. The libraries were normalized and pooled formultiplexed sequencing on an Illumina Hiseq4000 sequencer (Illumina, San Diego,CA, USA) using the pair-end 150bp run format (300 cycle run).