RNA-seq of argillite enrichment culture for archaea with medium R2A A + reducing agents in serum bottle at 36 deg
From the enrichment cultures, selected DNA samples were subjected to PCR amplification of archaeal 16S rRNA genes; the resulting amplicon library was purified from the reaction components by agarose gel electrophoresis, separated from the gel, and the concentrated samples were pooled for subsequent sequencing at Microsynth AG (Switzerland) using the second generation 454 sequencing technology. The amplicon library was constructed using two-part PCR, in which the DNA sequences were amplified, and the sorting tag and GS Titanium adapter sequences required for pyrosequencing were added. Multiple step PCR was used to attach GS Titanium Shotgun adaptor sequences. Region specific primers Arch349F and A934b (Grosskopf, Janssen, & Liesack, 1998); (Takai & Horikoshi, 2000) were used to target 16S rRNA V3-V5 genes region of archaea.