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R2C2+UMI: Combining concatemeric consensus sequencing with unique molecular identifiers enables ultra-accurate sequencing of amplicons on Oxford Nanopore Technologies sequencers.

ABSTRACT: The sequencing of PCR amplicons is a core application of high-throughput sequencing technology. Using unique molecular identifiers (UMIs), individual amplified molecules can be sequenced to very high accuracy on an Illumina sequencer. However, Illumina sequencers have limited read length and are therefore restricted to sequencing amplicons shorter than 600bp unless using inefficient synthetic long-read approaches. Native long-read sequencers from Pacific Biosciences and Oxford Nanopore Technologies can, using consensus read approaches, match or exceed Illumina quality while achieving much longer read lengths. Using a circularization-based concatemeric consensus sequencing approach (R2C2) paired with UMIs (R2C2+UMI) we show that we can sequence ~550nt antibody heavy-chain (IGH) and ~1500nt 16S amplicons at accuracies up to and exceeding Q50 (<1 error in 100,0000 sequenced bases), which exceeds accuracies of UMI-supported Illumina paired sequencing as well as synthetic long-read approaches.

SUBMITTER: Deng DZQ

PROVIDER: S-EPMC10473586 | biostudies-literature | 2023 Aug

REPOSITORIES: biostudies-literature

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R2C2+UMI: Combining concatemeric consensus sequencing with unique molecular identifiers enables ultra-accurate sequencing of amplicons on Oxford Nanopore Technologies sequencers.

Deng Dori Z Q DZQ Verhage Jack J Neudorf Celine C Corbett-Detig Russell R Mekonen Honey H Castaldi Peter J PJ Vollmers Christopher C

bioRxiv : the preprint server for biology 20230821

The sequencing of PCR amplicons is a core application of high-throughput sequencing technology. Using unique molecular identifiers (UMIs), individual amplified molecules can be sequenced to very high accuracy on an Illumina sequencer. However, Illumina sequencers have limited read length and are therefore restricted to sequencing amplicons shorter than 600bp unless using inefficient synthetic long-read approaches. Native long-read sequencers from Pacific Biosciences and Oxford Nanopore Technolog ...[more]

PMID: 37662385

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Dataset Information

R2C2+UMI: Combining concatemeric consensus sequencing with unique molecular identifiers enables ultra-accurate sequencing of amplicons on Oxford Nanopore Technologies sequencers.

Publications

R2C2+UMI: Combining concatemeric consensus sequencing with unique molecular identifiers enables ultra-accurate sequencing of amplicons on Oxford Nanopore Technologies sequencers.

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