title: 16S of Intercropping:monocropping and intercropping fields
design: Total genomic DNA was extracted from samples with a DNA isolation kit (MoBio, USA) following the manufacturers protocol and was stored at 80 C for further analysis. The 341F (5-CCTACGGGNGGCWGCAG-3) and 805R (5-GACTACHVGGGTATCTAATCC-3) primers were  used to amplify the bacteria-specific V3V4 hypervariable region of the 16S rRNA gene. Each primer contained 6-bp-long specific barcode sequences at the 5 end. All PCR reactions were carried out under  the following conditions: initial denaturation at 95 C for 3 min, followed by 25 cycles of denaturation at 95 C for 30 s, annealing at 55 C for 30 s, and elongation at 72 C for 30 s, with a final hold at 16 C for 2 min. PCR products were mixed in equal density ratios. Then the mixture of PCR products was run on 2% (w/v) agarose gels and extracted with the GeneJET Gel Extraction Kit (Thermo Scientific). Sequencing libraries were generated using NEB Next Ultra DNA Library Prep Kit forIllumina (NEB, USA) following the manufacturers recommendations and the sequencing was performed on the MiSeq platform.