16S rRNA gene profiling using micelle PCR approach of sludge sample 1
The protocol utilized a two-step micelle PCR (micPCR) protocol. In the first step, micPCR was performed using modified 357F and 926R primers that amplified the V3-V5 regions of 16S rRNA genes and which incorporated universal sequence tails at their 5Őends. In the second step, a micPCR was again used, but to amplify micPCR amplicons obtained from the first step micPCR. The second step micPCR utilized primers containing complementary sequences to the universal tails and included additional 454 sequencing-specific nucleotides, and specimen-specific MIDs. For both amplification steps, water-in-oil emulsions were prepared using the Micellula DNA Emulsion Kit (Roboklon).