Amplicon sequencing: Wheat and Soybean root microbiomes
The V4 hyper variable region of the 16s rRNA gene was amplified using the F515-R806 primers. Samples were amplified using two rounds of PCR, following the procedures detailed in Rascovan et al. 2013. The first round was performed to amplify the V4 hyper variable region of the 16s rRNA (30 cycles) and a second round was performed to add barcodes (10 cycles). Duplicated reactions were performed in both rounds of PCR to reduce PCR biases. All amplicons were cleaned using Ampure DNA capture beads (Agencourt- Beckman Coulter, Inc.) and pooled in equimolar concentrations before sequencing on a Genome Sequencer FLX (454-Roche Applied Sciences) using Titanium Chemistry according to the manufacturerâs instructions.