Figure 1.

An overview of RiboFR-Seq method. (A) A schematic diagram shows the process for capturing both ribosomal RNA variable regions and their flanking sequences simultaneously. (B) Experimental flowchart of RiboFR-Seq. Briefly, restriction enzymes with a 6-bp recognition site that can cleave bacterial 16S rRNA sequences were used to digest metagenomic DNA. Subsequently, the enzyme-digested DNA fragments with sticky end were self-circularized by direct intra-molecule ligation, which were then used as templates for LD-IPCR with specific inverse primers (labeled with biotin). LD-IPCR products were fragmented, of which contained biotin could be collected by magnetic streptavidin beads, then constructed NGS library for high- throughput sequencing.