Award details

minoTour: A real time analysis and data management platform for Oxford Nanopore minION reads.

Reference	BB/M020061/1
Principal Investigator / Supervisor	Professor Matthew Loose
Co-Investigators / Co-Supervisors
Institution	University of Nottingham
Department	School of Life Sciences
Funding type	Research
Value (£)	147,769
Status	Completed
Type	Research Grant
Start date	01/07/2015
End date	31/01/2017
Duration	19 months

Abstract

Amongst the many benefits of nanopore based sequencing approaches is the real-time generation of sequence-data. This can be exploited in many significant ways, for example detection of pathogens, monitoring sequence breadth and depth or even simply sequencing until a specific event is detected. I have developed a suite of tools as proof-of-principle for such approaches, called minoTour, which already demonstrates many of the benefits of real-time analysis. These tools require a database to store key read metrics in (currently mySQL based for speed of development) and a web based front end for data summaries and analysis. We propose developing this tool into a complete set of applications which can be used by individual users of minIONs or by centralised facilities wishing to support multiple minION users. We will also explore the provision of a centralised platform for real-time minION analysis and archival of data in conjunction with a service such as DNANexus. The system would be modular in design allowing key steps within it to be changed depending on the application, for example choosing a specific alignment tool for a given application. A number of groups are in the process of developing tools for the minIONs and long read technology in general. We seek to develop a standard for the archiving of minION data such that it can be readily manipulated by a range of tools as they are developed by the wider community. This standardisation will also allow for the development of compression standards for minION data. minoTour is already being exploited by members of the minION access program and its development is being observed by Oxford Nanopore Technologies. We will use the feedback generated from this in directing our work over the course of the grant.

Summary

The last 15 years has seen a dramatic explosion in the impact that DNA sequencing has had on our lives. From the announcement of the first draft human genome sequence in 2000 at an estimated cost of $2.7 billion to the announcement of the $1000 genome in 2014, the rate of growth in sequencing technology has been exponential. The main drive behind the dramatic lowering in cost is the development of massively parallel short read sequencing technologies from companies such as Illumina. These approaches generate short fragments of sequence, typically 75 - 300 base pairs in length, from the material being sequenced which are subsequently combined computationally to reassemble the original sequence. These approaches have generated vast quantities of data and revealed much that we did not know about biology. The establishment of long read sequencing platforms, first developed by PacBio, once again hold the potential for a step change in sequencing technologies. In particular, the portable nanopore based sequencer, minION, developed by Oxford Nanopore Technologies represents a significant new technology. In contrast with previous technologies, sequence reads are generated in almost real-time. As these reads can be very long they can provide information about the material being sequenced almost immediately. Currently no tools exist which truly exploit these real-time features. We are participating in the minION access program and have developed the first real-time analysis platform, as a proof of principle, called minoTour. This suite of tools aims to rapidly identify DNA sequences as they are generated and report the results back to the experimentalist via a simple web-based interface. Key components include sending notifications to an experimentalist in real-time as events take place. For example the detection of a sequence from a specific pathogen or achieving enough sequence coverage to recognise key events from sequence data. The key advantages of a web-based interfaceto such a platform are ease of use for an experimentalist in a wide variety of environments. Currently the minION platform requires an always on connection to the web for generation of the sequence data. In the future it is anticipated that this technology will be available on the local machine. The tools we are developing will also be able to run without a network connection allowing experimentalists to sequence and analyse data in the field. A broader extension of our approach focusses on de novo assembly of sequence data as it is generated. Early results suggest that de novo assembly from minION data is possible and this would allow for another broad range of applications both in the laboratory and the field. In the future tools such as the minION will be developed as basic sensing devices used in a number of different environments. Having real-time access to the data they generate in a manner similar to that proposed here would allow for the rapid detection of contaminants in a wide range of environments. Similarly it is easy to imagine the lab scientist wishing to sequence a DNA construct having their results within 30 minutes, representing significant savings in time and effort for a large number of projects.

Impact Summary

Over the last ten years the scale of sequencing achievable within individual laboratories has shifted from single plasmids to whole genomes. The establishment of next generation sequencing is key to answering many of the questions driving biology today. The recent rapid developments in the field of portable nanopore sequencing devices opens a vast range of opportunities. The tools that we develop for this technology are likely to impact research, researchers and industry in a wide range of disciplines. Rapid real time analysis of single molecule sequencing will reduce consumables costs, enable rapid sequence identification and ultimately allow a whole new class of environmental sensing. We anticipate significant rapid benefits in fields such as microbiology and the identification of pathogens, experiments focussed on phasing and requiring long reads, de novo assembly of small genomes and other approaches requiring long reads. We also anticipate developments which will translate into wider industry through our interactions with Oxford Nanopore Technologies themselves and other interested parties. Other sequencing companies are working towards long read sequence technology and are expressing interest in the approaches we are presenting. The development of minoTour will also have an impact on: a. Researchers: Through the acquisition of formalized training in sequence analysis and real time data processing. These will include both researchers within the UoN, partners collaborating on this proposal and members of the wider Midlands Sequencing Consortium, of which Deep Seq is a founding member, and further afield. b. The University of Nottingham: Developing novel sequencing tools at Nottingham will further support the centre in making a unique contribution to sequencing activities in the Midlands and beyond. c. The Midlands: Deep Seq is a member of the midlands sequencing consortium, which exists to share resources with compatible sequencing platforms across multiple Midlands Universities including Sheffield and Leicester alongside the M5 grouping. The development of tools supporting novel sequencing approaches will be shared amongst this grouping. d. The international relationships between Nottingham University and researchers outside the UK through collaborations between Nottingham researchers and the international research community including international companies. The wider public will also benefit in the longer term from the research conducted at DeepSeq through the increased ability of different industrial sectors to respond to their customer needs from the environment, to agriculture and health. The research achievements from DeepSeq and the Midlands Sequencing Consortium will be communicated to a range of audiences via presentations through to discussions and workshops with industry contacts, publications in journals targeting a wide range of audiences and conferences. The research from DeepSeq and its potential will also be communicated to the general public through the yearly University of Nottingham May Fest and through the 'Nottingham Potential' outreach activities.

Committee	Research Committee C (Genes, development and STEM approaches to biology)
Research Topics	Technology and Methods Development
Research Priority	X – Research Priority information not available
Research Initiative	Tools and Resources Development Fund (TRDF) [2006-2015]
Funding Scheme	X – not Funded via a specific Funding Scheme

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