16S rRNA amplicon sequencing of wastewater
The V3-V4 regions of the 16S rRNA gene were amplified using primer pairs 338F (5'-ACTCCTACGGGAGGCAGCAG-3') and 806R (5'-GGACTACHVGGGTWTCTAAT-3'). The PCR amplifications were performed on a T100 Thermal Cycler PCR thermocycler (BIO-RAD, USA) under standard PCR conditions. The PCR product was extracted from 2% agarose gel, purified using the PCR Clean-Up Kit (YuHua, Shanghai, China), and quantified using Qubit 4.0 (Thermo Fisher Scientific, USA). Purified amplicons were pooled in equimolar concentrations and sequencing on an Illumina PE300/PE250 platform in paired-end mode.