Metagenome sequence of SIK wastewater
Attachment of dual indices and Illumina sequencing adapters was performed in a 50 l reaction mix consisting 5 l PCR product DNA, 5 l Illumina Nextera XT Index Primer 1 (N7xx), 5 l Nextera XT Index Primer 2 (S5xx), 25 l 2 KAPA HiFi HotStart Ready Mix, and 10 l PCRgrade water. The thermocycling conditions consisted of initial denaturation at 95 C for 3 min, followed by 8 cycles of 95 C for 30 s, 55 C for 30 s, and 72 C for 30 s, and a final extension at 72 C for 5 min before cooling to 4 C. Constructed 16S metagenomic libraries were purified with Agencourt AMPure XP beads and quantified with Quant-iT PicoGreen. The libraries were validated on Agilent Technologies 2100 Bioanalyzer to ascertain the quality and library size distribution. Libraries were normalized and pooled. Pooled libraries were denatured with NaOH, diluted with hybridization buffer to a final concentration of 10 pM and then heat-denatured before MiSeq sequencing. PhiX Control kit v3 (20%; Illumina) were added to each run as internal control.