Shotgun human fecal samples
Pair-end libraries were elaborated from 0.2ng/ml DNA, strands were enzymatically fragmented with ATM to 300bp-600bp and complemented with unique combinations of Nextera XT index (REF: 15055293/ LOT: 20231971) in a barcode sample specific fashion. PCR was carried out under the following conditions: initial denaturation for 3 min at 72C and 30 s at 95C, followed by 12 cycles of denaturation for 10 s at 95C, annealing for 30 s at 55C and elongation for 30 s at 72C, and a final elongation step for 5 min at 72C. The genomic amplification was recovered using magnetic pearls washed with 80% ethanol and H20 qPCR for dilution. PCR products were quantified using Qubit dsDNA BR/HS Assay kit and fragment length was assessed with Bioanalyzer Agilent Technology. A final library for sequencing was created with equimolar ratios of libraries from each sample.