Realbio Technology

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Introduction Pipeline Advantages Sample Typical Cases

16S rDNA encodes 16S ribosomal RNA and is present in all bacteria. The gene sequences contain several hypervariable regions that can provide species-specific signature sequences useful for identification of bacteria. In addition, 16S rRNA genes are used in reconstructing phylogenies, which is due to the slow rates of evolution of the genes. 16S rDNA sequencing is a common and well-established amplicon sequencing method for studying phylogeny and taxonomy of samples from complex microbiomes or environments that are challenging to study.

The internal transcribed spacers (ITS) region is the most widely sequenced DNA region in molecular ecology of eukaryotes and has been recommended as the universal fungal barcode sequence. There are two ITSs in eukaryotes, ITS1 is located between 18S and 5.8S rRNA genes, while ITS2 is between 5.8S and 26S rRNA genes. The ITS region is widely used in taxonomy and molecular phylogeny because of high degree of variation even between closely related species.

Research Field

1)Medicine: digestive system disease (Inflammatory Bowel Disease), endocrine system disease (obesity, diabetes), etc.
2)Agriculture: rhizosphere microorganisms, soil microorganisms treated by fertilization, etc.
3)Animal husbandry: animal stomach, intestines, etc.
4)Environment: soil, water, mud, fog, etc.
5)Special extreme environment: mining microorganisms, Yellowstone national park microorganisms, etc.

Sequencing Strategy
1)HiSeq 2500 platform
2)Paired-end 250 bp

Research Contents
1)Data quality control.
2)OTUs (Operational Taxonomic Units) clustering.
3)OTUs analysis and species annotation, including KRONA results, phylogenetic composition analysis, phylogenetic tree, heatmap, and taxonomic tree.
4)Alpha-diversity analysis, including rarefaction curve, rank abundance curve, and alpha indices table.
5)Beta-diversity analysis, including weighted/unweighted UniFrac distance heatmap, PCA, PCoA, UPGMA, and NMDS analysis.
6)Multi-variate statistical analysis, including LEfSe analysis, CCA/RDA, STAMP, and GraPhlAn analysis.