Nanopore takes over 16S rRNA gene amplicon sequencing
)
16S rRNA gene amplicon sequencing remains a popular and widely used method to study bacterial community in multiple environments, although its resolution in bacterial classification is somewhere between genus and species level. For over a decade 16S rRNA gene amplicon sequencing was a domain of next generation sequencing (NGS) platforms. NGS was first to allow for high throughput analysis of multiple samples despite their main limitation that was and still remains: a short read. To surpass this limitation only a short fragment of the full 16S rRNA gene was amplified and subjected for sequencing. This resulted in development of multiple protocols targeting different variable regions of 16S rDNA causing problem with data comparability across many studies. Nanopore sequencing is the only technology that allows high throughput, realtime sequencing of near-full-length 16S rRNA gene amplicons at low cost, yet still suffers from a relatively high error rate on a single molecule level. The utilisation of unique molecular identifiers (UMI) brought a hope for resolving this setback, but in rich bacterial communities would require ultra-deep sequencing what would have jeopardised its cost-effectiveness. It is clear that by fixing the problem related to primers universality (targeting more bacteria) and reducing the error on a single molecule level combined with low running costs, would make nanopore based sequencing technology the most obvious approach in 16S rRNA gene amplicon sequencing strategy.
