16S sequencing of positive control samples

Synthesized mock DNA samples were designed as positive control, named gene blocks. We selected DNA to synthesize using regions of the 16S rRNA gene in 8 archaeal species which would not normally be detected in experimental data because the sequences at the amplification primer binding sites in the archaeal V1V2 region do not match the bacterial V1V2 primers used. However, in the engineered sequences, correct bacterial 16S V1V2 primer bindings sites were added synthetically to archaeal controls, allowing amplification. This has the advantage that the output sequences can be worked up as for experimental samples and the archaeal sequences read out with no extra effort.

本研究设计合成模拟DNA样本作为阳性对照，将其命名为基因片段（gene blocks）。我们选取8种古菌的16S核糖体RNA基因（16S rRNA gene）区域进行DNA合成，这类古菌通常无法在实验数据中被检测到，原因是其V1V2区域的扩增引物结合位点序列与所使用的细菌V1V2引物不匹配。但在经工程化改造的序列中，我们通过合成手段为古菌对照添加了匹配的细菌16S V1V2引物结合位点，使其能够实现扩增。该设计的优势在于，输出序列可按照实验样本的标准流程进行处理，且无需额外操作即可直接读取古菌序列的相关信息。