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Highly accurate barcode and UMI error correction using dual nucleotide dimer blocks allows direct single-cell nanopore transcriptome sequencing

Martin Philpott, Jonathan Watson, Anjan Thakurta, Tom Brown Jr, Tom Brown Sr, Udo Oppermann, Adam P Cribbs
doi: https://doi.org/10.1101/2021.01.18.427145
Martin Philpott
1Botnar Research Centre, Nuffield Department of Orthopedics, Rheumatology and Musculoskeletal Sciences, National Institute of Health Research (NIHR) Oxford Biomedical Research Centre (BRC), University of Oxford, Oxford OX3 7DQ, United Kingdom
2Oxford Centre for Translational Myeloma Research Oxford, Oxford OX3 7DQ, United Kingdom
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Jonathan Watson
3Radcliffe Department of Medicine, Oxford University
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Anjan Thakurta
2Oxford Centre for Translational Myeloma Research Oxford, Oxford OX3 7DQ, United Kingdom
3Radcliffe Department of Medicine, Oxford University
4Bristol Myers Squibb, Translational Development and Diagnostics, Summit, NJ, USA
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Tom Brown Jr
5ATDBio Ltd, Building 30, School of Chemistry, University of Southampton, United Kingdom, SO17 1BJ
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Tom Brown Sr
5ATDBio Ltd, Building 30, School of Chemistry, University of Southampton, United Kingdom, SO17 1BJ
6Department of Chemistry, University of Oxford, Chemistry Research Laboratory, 12 Mansfield Road, Oxford, OX1 3TA, United Kingdom Structural Genomics Consortium, University of Oxford, Oxford OX3 7LD, United Kingdom
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Udo Oppermann
1Botnar Research Centre, Nuffield Department of Orthopedics, Rheumatology and Musculoskeletal Sciences, National Institute of Health Research (NIHR) Oxford Biomedical Research Centre (BRC), University of Oxford, Oxford OX3 7DQ, United Kingdom
2Oxford Centre for Translational Myeloma Research Oxford, Oxford OX3 7DQ, United Kingdom
7Centre for Medicines Development, University of Oxford
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  • For correspondence: adam.cribbs@ndorms.ox.ac.uk udo.oppermann@ndorms.ox.ac.uk
Adam P Cribbs
1Botnar Research Centre, Nuffield Department of Orthopedics, Rheumatology and Musculoskeletal Sciences, National Institute of Health Research (NIHR) Oxford Biomedical Research Centre (BRC), University of Oxford, Oxford OX3 7DQ, United Kingdom
2Oxford Centre for Translational Myeloma Research Oxford, Oxford OX3 7DQ, United Kingdom
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  • For correspondence: adam.cribbs@ndorms.ox.ac.uk udo.oppermann@ndorms.ox.ac.uk
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Abstract

Droplet-based single-cell sequencing techniques have provided unprecedented insight into cellular heterogeneities within tissues. However, these approaches only allow for the measurement of the distal parts of a transcript following short-read sequencing. Therefore, splicing and sequence diversity information is lost for the majority of the transcript. The application of long-read Nanopore sequencing to droplet-based methods is challenging because of the low base-calling accuracy currently associated with Nanopore sequencing. Although several approaches that use additional short-read sequencing to error-correct the barcode and UMI sequences have been developed, these techniques are limited by the requirement to sequence a library using both short- and long-read sequencing. Here we introduce a novel approach termed single-cell Barcode UMI Correction sequencing (scBUC-seq) to efficiently error-correct barcode and UMI oligonucleotide sequences synthesized by using blocks of dimeric nucleotides. The method can be applied to correct either short-read or long-read sequencing, thereby allowing users to recover more reads per cell and permits direct single-cell Nanopore sequencing for the first time. We illustrate our method by using species-mixing experiments to evaluate barcode assignment accuracy and evaluate differential isoform usage and fusion transcripts using myeloma and sarcoma cell line models.

Competing Interest Statement

A.T is a full-time employee and shareholder of Bristol Myers Squibb. JW, TBj and TBs are shareholders of ATDBio. MP, UO, A.P.C are inventors on patents filed by Oxford University Innovations for single-cell technologies.

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-ND 4.0 International license.
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Posted January 19, 2021.
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Highly accurate barcode and UMI error correction using dual nucleotide dimer blocks allows direct single-cell nanopore transcriptome sequencing
Martin Philpott, Jonathan Watson, Anjan Thakurta, Tom Brown Jr, Tom Brown Sr, Udo Oppermann, Adam P Cribbs
bioRxiv 2021.01.18.427145; doi: https://doi.org/10.1101/2021.01.18.427145
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Highly accurate barcode and UMI error correction using dual nucleotide dimer blocks allows direct single-cell nanopore transcriptome sequencing
Martin Philpott, Jonathan Watson, Anjan Thakurta, Tom Brown Jr, Tom Brown Sr, Udo Oppermann, Adam P Cribbs
bioRxiv 2021.01.18.427145; doi: https://doi.org/10.1101/2021.01.18.427145

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