TCE-degrading mixed culture
The SMRTbell library was prepared by following the procedure by Pacific Biosciences of California, Inc. for the amplification and sequencing of full-length 16S genes (Unknown, 2021). In brief, equal molar amounts of each barcoded PCR product were pooled, and 500-1000 ng of pooled amplicon sample was used for DNA damage repair followed by end-repair/A-tailing and ligation steps to introduce the universal hairpin adapters onto double-stranded DNA fragments. After purification with AMPure PB beads to remove the adapter dimer, the SMRTbell library was incubated with Sequencing Primer v4 and Sequel II Binding Kit 2.1 for primer annealing and polymerase binding. Finally, sequencing was performed in circular consensus sequence (CCS) mode on a PacBio Sequel IIe instrument to generate HiFi reads with predicted accuracy (Phred Quality Score) = 30 (Q30, equivalent to 99.9% base calling accuracy).