The third generation sequencing technology represented by the latest PacBio Sequel sequencing system adopts SMRT (single molecular real-time) technology, which can achieve single molecule level and long reads sequencing. It can easily read the full-length of rRNA genes / ITS sequence of microorganisms, no longer subject to the limitations of short reads. Additionally, PacBio's unique circular consensus sequencing mode (CCS) can greatly improve the accuracy of single base sequencing, far more than the accuracy of second-generation sequencing, which guarantees the accuracy of the obtained rRNA gene /ITS full-length sequence. Based on the third generation sequencing features, PacBio's SMRT sequencing technology can greatly improve our capacity to analyze microbiota diversity and compositional profiling in depth at the level of species and even strains.

Advantages

Principle of CCS sequencing mode

The sequence variations in V1-V9 regions of 16S rRNA gene are not consistent. The full-length sequence of 16S rRNA gene can reflect species-level information more comprehensively. Figure a, represented by Salmonella, shows 97.4% Interspecies differences in the full-length sequence, but the V4 region is highly conserved, and thus the second generation sequencing results will underestimate the diversity of this species; Figure b, the diversity of some species in V4 region may be higher than that in other V regions. The second generation sequencing results will overestimate the diversity of related species. *

With the comparison of sequencing results between two platforms, the results of third generation sequencing significantly reduced the uncertainty of species classification information.

Comparing the sequencing results of 16S rRNA genes between PacBio platform and Illumina platform, the results of PacBio platform can significantly improve the detection accuracy at species level, and greatly reduce the uncertainty of species classification.

Detailed composition profiles at species level

Flora composition heatmap at species level  

Violin diagram of abundance distribution at species level with significant difference between groups

Personalbio provides 16S rRNA gene & ITS full-length sequencing and analysis services to advance microbiome researches by the latest Sequel system. The power of SMRT technology, the age of precision science!