Evaluation of a Nucleic Acid Preservation Protocol for Microbiome Studies Involving Samples from the Oral Cavity
Total bacterial DNA was isolated from all samples. After DNA concentration measurements, DNA was used as input for PCR amplification of the 16S rRNA region. The Illumina 16S rRNA Primer Constructs (515F-806R) for the amplification were fused with Golay indices and adapter sequences as described in the Earth Microbiome Project protocol (http://www.earthmicrobiome.org/protocols-and-standards/16S/). PCR products were purified and quantified. The libraries were equimolarly pooled and prepared for Illumina sequencing. Sequencing was performed on a MiSeq apparatus (Illumina) with 251 cycles. After sequencing, raw data was demultiplexed and further denoised by using DADA2  pipeline.