RT Journal Article
SR Electronic
T1 Highly accurate barcode and UMI error correction using dual nucleotide dimer blocks allows direct single-cell nanopore transcriptome sequencing
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 2021.01.18.427145
DO 10.1101/2021.01.18.427145
A1 Martin Philpott
A1 Jonathan Watson
A1 Anjan Thakurta
A1 Tom Brown, Jr
A1 Tom Brown, Sr
A1 Udo Oppermann
A1 Adam P Cribbs
YR 2021
UL http://biorxiv.org/content/early/2021/01/19/2021.01.18.427145.abstract
AB 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 StatementA.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.