Metabarcoding of Wood from flooded tree, tropical reservoir
Diversity analysis was carried out on active (cDNA-converted from RNA) communities. The V3-V4 region of the 16S rRNA gene transcripts was amplified by PCR using the universal primer set 341F - 806R for bacteria and archaea. Each PCR reaction (50 L final volume) contained 1.25 U of TaKaRa PrimeSTAR GXL DNA polymerase (OZYME), 0.4 M of each primer, and 5 L of previously diluted cDNA template (1/5 and 1/10 dilution factors for wood and biofilm samples, respectively). The correct amplicon size and purity was checked by agarose gel electrophoresis. The 16S amplicons were sequenced on MiSeq Illumina technology (paired end 2 x 250 bp) at the DNA Sequencing Facility of the Madison Biotechnology Center, University of Wisconsin.