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Method Article
UMI-linked nanopore consensus sequencing (UMIC-seq) of highly similar gene variants
Florian Hollfelder
University of Cambridge
Corresponding Author
ORCiD: https://orcid.org/0000-0002-1367-6312
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Here we present a straightforward unique molecular identifier (UMI)-linked nanopore consensus sequencing workflow (UMIC-seq), resulting in cost-effective and accurate long-read sequencing of amplicons. Short random molecular barcodes (i.e. unique molecular identifiers, UMIs) are attached to a pool of gene variants prior to nanopore sequencing to enable reliable clustering and the generation of accurate consensus sequences, even when starting from highly similar gene variants (e.g. a library of point mutants in directed protein evolution) that could not be reliably distinguished in the ordinary nanopore sequencing output.
Keywords
nanopore sequencing, consensus sequences, amplicon sequencing, directed evolution, protein engineering
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Subject Areas
Biochemistry
Molecular biology
Biological techniques
Computational biology and bioinformatics
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This is a preprint. It has not completed peer review.
Method Article
UMI-linked nanopore consensus sequencing (UMIC-seq) of highly similar gene variants
Florian Hollfelder
University of Cambridge
Corresponding Author
ORCiD: https://orcid.org/0000-0002-1367-6312
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
History
CURRENT STATUS: Posted
Version 1
Posted 30 Nov, 2020
No community comments so far
Metrics
Comments: 0
HTML Views: ...
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Here we present a straightforward unique molecular identifier (UMI)-linked nanopore consensus sequencing workflow (UMIC-seq), resulting in cost-effective and accurate long-read sequencing of amplicons. Short random molecular barcodes (i.e. unique molecular identifiers, UMIs) are attached to a pool of gene variants prior to nanopore sequencing to enable reliable clustering and the generation of accurate consensus sequences, even when starting from highly similar gene variants (e.g. a library of point mutants in directed protein evolution) that could not be reliably distinguished in the ordinary nanopore sequencing output.