Mining data-rich reads with Philipp Rescheneder

Having the right tools for the job is key for success. For the data-rich, any-length reads produced by nanopore sequencing, the right bioinformatics tools for analysis should be able to parse out the full scope of variants and epigenetic information, as well as phase the reads.

Philipp Rescheneder (Senior Director of Application Bioinformatics at Oxford Nanopore Technologies) and his team identify the best existing, as well as create new, analysis tools and pipelines for DNA and RNA nanopore sequencing data. Find out how their work helps you to maximise the insights you can gain from your nanopore datasets.

Creating and optimising tools for nanopore reads

When it became clear that having access to increased read lengths enabled interrogation of repetitive regions in the genome, there was a buzz at Cold Spring Harbor Laboratory due to the potential to study structural variants (SVs). However, Michael Schatz and his team very quickly realised that the bioinformatics tools available, which had all been optimised to work with short-read sequencing data, could not answer their questions.

To address this, Philipp, a visiting PhD student at the time, developed an alignment tool optimised for structural variant analysis and datasets containing reads of any length, whilst his colleague Fritz Sedlazeck worked on an SV caller called Sniffles. Since then, Sniffles, and now Sniffles2, has been widely used for analysing SVs in nanopore data, and Fritz’s and Philipp’s working relationship still holds today. You can hear more from Fritz by viewing his talk from NCM Singapore 2023.

This experience has fed into Philipp’s role at Oxford Nanopore. The foundation of his team’s work is benchmarking: testing new and improved tools against truth sets so that they can define what is best practice for nanopore data analysis. When they identify gaps, the team initiate the development of a new tool or look to experts external researchers for collaboration.

‘… that's where collaborations, I think, come in and work really well. We can work with somebody who has core expertise in a certain field, and we can supplement that with nanopore-specific knowledge’

Combining the best tools for a unified approach

Benchmarking and tool development have bought the analysis of nanopore data a long way. However, as Oxford Nanopore data gives you the enticing ability to understand a range of variant types, all from one sequencing run, in Philipp’s mind, the next challenge is to be able to assess all of them together in one easy workflow.

Historically, prior to nanopore sequencing, researchers could only assess one or a small number of variant types with the sequencing technologies that were available. This meant that bioinformaticians developed tools that were optimised to assess one variant type, such as single nucleotide variants, SVs, copy number variants, methylation variants. Therefore, now, when researchers want to investigate the full scope of genetic variation, the tools are siloed.

‘So what happens is you have this ecosystem of lots of tools that look at very specific kinds of things. But they all run separately.’

Biologically and technically, these variants could be linked, and together they would form a complete picture of the genetic variation present, but legacy has made it difficult to put these analyses together to see how variants interact. This obstacle drives Philipp and his team to work on cohesive and unified approaches for genomic analysis.

Striving for a holistic understanding of biology

One such approach is to feed insights from the benchmarking and collaborations with researchers to the EPI2ME team. EPI2ME is an application providing a range of simple-to-use bioinformatic workflows, where the best tools are consolidated to seamlessly provide you with all the information required for a complete understanding of the underlying biology.

These workflows are curated and maintained by nanopore data analysis experts, so rather than developing your own pipelines, you can trust EPI2ME to deliver accurate and high-performing analyses targeted at specific research questions.

For example, with genomic instability being a hallmark of cancer, a holistic assessment of variation could improve our understanding of both the mechanisms driving the accumulation of variants, and how these variants combine into the resulting cancerous phenotype.

‘…if you have all of this, you can not only understand what’s going on, but also how it happened. Then, potentially, you could use this information for early detection or even drug targets.’

Oxford Nanopore’s EPI2ME somatic variation workflow helps researchers take this approach by detecting single nucleotide variants, complex SVs, and modifications, and phasing haplotypes, in tumour-normal sample sets. Visit the somatic variation workflow poster to learn more.

One of the tools in the pipeline, Severus, was developed by Mikhail Kolmogorov’s group (National Institute for Health, US) and supported by a member of Philipp’s team through a collaboration. It was created as most existing tools were designed for the analysis of germline variation, limiting our insights and revealing only part of the story.

The somatic variation workflow is only one of the many pipelines that the EPI2ME team and Philipp’s team have worked on. You can find a workflow tailored to your research by seeing the full range of workflows and downloading the software from the EPI2ME website.

So what does the future hold?

Looking back, for Philipp, it has been fantastic to observe the growth of analysis tools developed for nanopore data, and see the unique ways that people are making use of the additional information. Going forwards, Philipp is particularly excited to see the breadth of insights uncovered when nanopore sequencing is applied to more population-wide studies.

Beyond this, Philipp is also driven by the potential to maximise access to genetic sequencing.

‘It [Oxford Nanopore] feels like the only technology that will be able to effect everyday life in a positive manner, just because it is portable, it is small. You have sequencers that you can put in your pocket, you can put it on a desk. It just opens up things that are not possible with a huge box somewhere in the lab … That is actually what’s the motivating factor of working at nanopore, being part of this development, seeing what implications it will have.’

If you want to hear more about the latest projects from Philipp, please come and join us at NCM Boston where he will be presenting the latest from his team. Don’t risk missing out!