DNase Max Kit (50)

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Nucleic Acid Extraction, Amplicon Library Preparation and Sequencing: The total RNA and DNA were co-extracted from 2 g of soil for each of the 60 samples using the RNeasy PowerSoil Total RNA kit (Qiagen, Hilden, Germany) with the RNeasy PowerSoil DNA Elution kit (Qiagen, Hilden, Germany), following the manufacturer’s instructions (Figure 1). After eluting the RNA samples from the capture column of the RNeasy PowerSoil Total RNA kit, the bound DNA on the capture column was further eluted using the RNeasy PowerSoil DNA Elution kit. Remaining DNA in RNA samples was removed using the DNase Max kit (Qiagen, Hilden, Germany). We performed PCR reactions for 10% of RNA samples to verify whether the amount of DNase applied was enough to ensure DNA was completely removed from RNA samples. The DNA-free RNA was converted to cDNA by incubating with random hexamers using the Transcriptor High Fidelity cDNA Synthesis Kit (Roche, Basel, Switzerland), which was then purified using the MinElute PCR Purification Kit (Qiagen, Hilden, Germany). DNA and cDNA were quantified using a NanoDrop 2000 Spectrophotometer (Thermo Fisher scientific, Waltham, MA, USA).

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