title: 16S_ZH15_vinerow_surface
design: DNA was extracted from 0.2 g of lyophilized soil using the NucleoSpin Soil Kit (Macherey-Nagel, Dren, Germany), following the manufacturers protocol with a duplicate extraction step to achieve more stable DNA recovery. DNA concentrations were determined using a Cary Eclipse fluorescence spectrophotometer (Agilent Technologies, Santa Clara, CA, USA) and normalized to 1 ng/L. To profile the prokaryotic community, the V3V4 regions of the 16S rRNA gene (hereafter "16S") was amplified using primers 341F and 805R. For fungal community analysis, the ITS2 region (hereafter "ITS") was amplified using a forward primer mix (ITS3ngsmix15) and the reverse primer ITS4ngsUni. All primers were extended with additional linkers, a frame shift to enhance sequence diversity, and Nextera adapter sequences for downstream indexing. The 16S PCR protocol included an initial denaturation at 95C for 3 minutes, followed by 20 cycles of denaturation at 98C for 20 seconds, annealing at 52C for 30 seconds, and extension at 72C for 30 seconds, with a final extension at 72C for 2 minutes. For ITS amplification, the PCR protocol was slightly different with 30 cycles of denaturation at 98C for 20 seconds, annealing at 55C for 20 seconds, and extension at 72C for 30 seconds. Each 10-L PCR reaction contained 1 KAPA HiFi HotStart DNA Polymerase ReadyMix (Roche, Basel, Switzerland), 0.5 M of each primer, and either 2 ng (16S) or 1 ng (ITS) of template DNA. PCR reactions were performed in triplicate and pooled. Amplicons were visualized on 1% agarose gels and purified using custom-made SPRI beads (GDC, Zurich, Switzerland). Indexing PCR was performed using the following conditions: initial denaturation at 95C for 3 minutes, followed by either 10 cycles (16S) or 5 cycles (ITS) of 95C for 30 seconds, 55C for 30 seconds, and 72C for 30 seconds, with a final extension at 72C for 5 minutes. Each 20-L indexing reaction consisted of 1 KAPA HiFi HotStart DNA Polymerase ReadyMix, 2 L each of Nextera XT Index primers 1 and 2 (Illumina, San Diego, CA, USA; Set A and D), 1 L dimethyl sulfoxide, and 4 L of purified PCR product. Following a second round of SPRI bead purification, DNA concentrations were quantified using the Qubit dsDNA BR Assay Kit (Thermo Fisher Scientific, Waltham, MA, USA). Equimolar amounts of each sample were pooled to construct separate 16S and ITS libraries. Libraries were size-selected by gel electrophoresis on a 1% agarose gel, excised, and purified using the NucleoSpin Gel and PCR Clean-up Kit (Macherey-Nagel), according to the manufacturers protocol. Final library quantification was performed using the QuDye dsDNA HS Kit (Lumiprobe, Hannover, Germany), and libraries were diluted to a final concentration of 4 nM. Sequencing was conducted on an Illumina MiSeq platform using a 600-cycle MiSeq Reagent Kit v3 (Cat. No. MS-102-3003), generating paired-end reads (2  300 bp). Library preparation and sequencing were carried out at the Genetic Diversity Centre, ETH Zurich (https://gdc.ethz.ch/).