B1_C06_wk8_mNaChBac
Bacterial genomic DNA was extracted using MO BIO PowerSoil DNA Isolation Kit (MO BIO Laboratories). The 16S rRNA V4-V5 region was amplified by PCR using the original Caporaso et al primers(Caporaso, Lauber et al. 2011) (515F: GTGCCAGCMGCCGCGGTAA; 806R: GGACTACHVGGGTWTCTAAT) and sequenced on the MiSeq platform (Illumina) using the 2 x 250 bp paired-end protocol.The 16S rRNA amplicon read pairs were demultiplexed and reads were merged using USEARCH v7.0.1090(Edgar 2010), allowing zero mismatches and a minimum overlap of 50 bases. Merged reads were trimmed at first base with Q5. A quality filter was applied to the resulting merged reads. Reads containing above 0.05 expected errors were discarded. 16S rRNA gene sequences were clustered into Operational Taxonomic Units (OTUs) at a similarity cutoff value of 97% using the UPARSE algorithm(Edgar 2013). OTUs were mapped to an optimized version of the SILVA Database(Quast, Pruesse et al. 2012), containing only the 16S V4 region to determine taxonomies. Abundances were recovered by mapping the demultiplexed reads to the UPARSE OTUs.