Sensitivity and correlation of hypervariable regions in 16S rRNA genes in phylogenetic analysis. Use of 16S rRNA and rpoB genes as molecular markers for microbial ecology studies. Case R J, Boucher Y, Dahllöf I, et al.Metagenomic survey of bacterial populations.Identification and taxonomic classification of bacterial species.PlatformĪdvantages and applications of 16S rRNA sequencing The common 16S rRNA sequencing regions by various platforms. The former three platforms only cover 100 to 600 base pairs per single read, while the Pacific Bioscience can produce full-length 16S rRNA. The common sequencing platforms for 16S rRNA sequencing include Illumina MiSeq, Roche 454, Illumina MiSeq/HiSeq, and Pacific Bioscience. The workflow of 16S rRNA sequencing for human microbiome analysis.ġ6S rRNA sequencing generally uses the next/third generation sequencing technology to read the PCR products which are amplified with suitable universal primers of one or several regions of 16S rRNA. It has been widely applied in basic research, as well as medical, forensic, agricultural, and industrial microbiology.įigure 3. 16S rRNA sequencing is featured by fast speed, cost-efficiency, and high-precision. Since 16S rRNA gene is conserved in bacteria, and contain hypervariable regions that can provide species-specific signature sequences, 16S rRNA sequencing is widely used in identification of bacteria and phylogenetic studies. Illustration of different variable regions (Yang et al. Green regions (V4, V5, and V6) are associated with the shortest geodesic distance, which implies that they may be the optimal choice for phylogeny-related analyses, including phylogenetic analysis of novel bacteria phyla.įigure 2. (Figure 2), red regions (V2 and V8) have a poor phylogenetic resolution at the phylum level. The quality of the DNA li-brary was examined using the Quibit® 3.0 Fluorometer 2.0 (Thermo Fisher Scientific, USA) and Agilent Bioana-lyzer 2100 system. Fox pioneered the use of 16S rRNA in phylogenetics. ucts were used to construct a DNA sequencing library using the the NEB Next® Ultra DNA Library Prep Kit from Illumina (San Diego, CA, USA) according to the manufacturer’s instruction. (C) The regions and primers of 16S rRNA.ġ6S rRNA gene is characterized by ubiquity and evolutionary properties, which allow it to become an important molecular marker in microbial ecology. Individual regions are shown in the same color (Yang et al. (B) The 2D and 3D structure of the 16S rRNA gene. (A) The involvement of 16S rRNA gene in a prokaryotic ribosome. You can find 16S rRNA sequences in databases such as Ribosomal database project (RDP), Greengenes database, Silva, and Human microbiome project (HMP). The bacterial 16S rRNA gene contains nine hypervariable regions (V1-V9) ranging from 30-100 base pairs, flanked by conserved regions (Figure 1C). Figure 1A shows how 16S rRNA is involved in a prokaryotic ribosome. 16 S ribosomal RNA (or 16 S rRNA) is the component of the 30S small subunit of a prokaryotic ribosome, roughly 1500 base pairs.
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