Capturing the dynamics of genome replication on individual ultra-long nanopore sequence reads

Journal article

Replication of eukaryotic genomes is highly stochastic, making it difficult to determine the replication dynamics of individual molecules with existing methods. We report a sequencing method for the measurement of replication fork movement on single molecules by detecting nucleotide analog signal currents on extremely long nanopore traces (D-NAscent). Using this method, we detect 5-bromodeoxyuridine (BrdU) incorporated by Saccharomyces cerevisiae to reveal, at a genomic scale and on single molecules, the DNA sequences replicated during a pulse-labeling period. Under conditions of limiting BrdU concentration, D-NAscent detects the differences in BrdU incorporation frequency across individual molecules to reveal the location of active replication origins, fork direction, termination sites, and fork pausing/stalling events. We used sequencing reads of 20–160 kilobases to generate a whole-genome single-molecule map of DNA replication dynamics and discover a class of low-frequency stochastic origins in budding yeast. The D-NAscent software is available at https://github.com/MBoemo/DNAscent.git.

Authors: Müller, C; Boemo, M; Spingardi, P; Kessler, B; Kriaucionis, S

• Publication status: Published
• Peer review status: Peer reviewed
• Publisher: Springer Nature
• Journal: Nature Methods
• Volume: 16
• Pages: 429-436
• Publication date: 2019-04-22
• Acceptance date: 2019-03-18
• DOI: 10.1038/s41592-019-0394-y
• EISSN: 1548-7105
• ISSN: 1548-7091
• Language: English
• Keywords: FFR