Marcus Stoiber: Detecting methylation and more from raw nanopore signal

Marcus Stoiber gallantly stepped into the breach to discuss the latest developments in modified base calling. He opened his presentation by describing Tombo and a series of improvements that have recently been implemented. The Tombo analysis platform utilises raw signal analysis and basecalled reads to allow detection of epigenetic modifications from raw nanopore sequencing data. Three different ways of detecting modified bases are available, providing users with flexibility with regard to application and experimental design.

Marcus explained that recent work on the generation of a number of motif-specific models (in human and E. coli) has dramatically improved the power of the approach and delivers more accurate calls. He also highlighted the availability of online tutorials that allow researchers to train the tool to detect base modifications in their organism of interest – providing it has a specific sequence motif/pattern. A further improvement to the tool is the introduction of ‘level sample comparison’ which allows improved modification detection in specific contexts, such as highly modified direct RNA.

In the next section of his talk, Marcus touched on the ‘flip-flop’ algorithm that Clive Brown revealed in his plenary presentation on day 1 of the meeting. The flip-flop algorithm simplifies base calling by outputting single bases as opposed to more complex k-mer labels. This provides a better architecture for calling base modifications, allowing the output to scale gently with the addition of different bases. Training the model is much easier and requires less data. He also provided data showing how flip-flow successfully called 5hmC with no loss in accuracy to the underling base.