Long-Term Antibiotic Treatment Does Not Increase Antibiotic Resistance in the Upper Airway or Gut Microbiome of Cystic Fibrosis
Total bacterial DNA was isolated from all samples. After DNA concentration measurements, DNA was used as input for PCR amplification of the V4 region of the rRNA cluster genes. The Illumina V4 Primer Constructs for the amplification were fused with Golay indices and adapter sequences as described in the Earth Microbiome Project protocol (http://www.earthmicrobiome.org/protocols-and-standards/16S/). PCR products were purified and quantified. The libraries were equimolarly pooled and prepared for Illumina sequencing. Sequencing was performed on a MiSeq apparatus (Illumina) with 251 cycles. After sequencing, raw data was demultiplexed and further denoised by using the DADA2  pipeline.