Illumina 16S metabarcoding of DNA-3RZ-SEM
SEM was prepared with a combination of 300 g of soil and 300 mL of distilled water homogenized by stirring for 4 hours. Subsequently, the mixture was centrifuged at 9,000 rpm for 10 minutes, the supernatant was recovered, and 1.5% agar was added, autoclaved, and plated in petri dishes. To obtain the culturable microbiota from the BS and RZ samples, a suspension of 1 g of soil in 1 mL of 1X Phosphate-Buffered Saline (PBS 1X) was used as inoculum. The mixture was homogenized by shaking in the Revolver Rotator (LabNet) for 2 hours and then decanted by gravity for 30 minutes at room temperature. 100 uL of supernatant were inoculated into SEM plates and distributed in the petri dish with an L-shaped cell Spreader. Subsequently, the plates were incubated at 30C for 72 h, and after this time, the bacterial growth was resuspended in 200 uL of 1X PBS to perform DNA isolation using the DNeasy Blood & Tissue kit (Qiagen), following the manufacturer's recommendations. The V1-V3 region of the bacterial 16S rRNA gene was amplified with the primers 28F (5-GAGTTTGATCNTGGCTCAG-3) and 519R (5-GTNTTACNGCGGCKGCTG-3). The amplicons were sequenced with an Illumina sequencer using the service of Molecular Research LP. (Mr. DNA Lab). In the case of the RNA samples, reverse transcription was performed prior to amplification to generate cDNA.