Diclofenac removal in optimized bach reactors under denitrifying conditions
Genomic DNA from the samples was extracted using the FastDNA SPIN Kit for Soil DNA Extraction (MP Biomedicals) following the manufacturer's recommendations. The sequencing and bioinformatics services were performed by the company GenomeDX  Advanced Genetics (Rio de Janeiro, RJ). In the PCR, the primer set 341F - 806R (Yu et al., 2005) of the 16S rRNA gene, variable region V3-V4 was used. PCR reactions were performed with Phusion High-Fidelity PCR Master Mix (New England Biolabs). Sequencing libraries were generated using an UltraNBNext Kit DNA library for Illumina (New England Biolabs, Ipswich, USA), following the manufacturer's recommendations. Sequences were assembled and filtered (QPhred ? 33) to remove sequences containing low quality bases. Chimeras were detected (UCHIME algorithm) and excluded. A sequence analysis was performed using the Uparse software (Uparse v7.0.1001 ? http: //drive5.com/uparse/) [6]. Sequences with similarity ?97% were assigned to the same OTUs (Operational Taxonomic Units). For each representative sequence, the Mothur software was used in comparison with the SSUrRNA database of the SILVA database (http://www.arb-silva.de/) (Wang et al., 2007).