Single molecule-level detection and long read-based phasing of epigenetic variations in bacterial methylomes

Beaulaurier, John; Zhang, Xue Song; Zhu, Shijia; Sebra, Robert; Rosenbluh, Chaggai; Deikus, Gintaras; Shen, Nan; Múnera, Diana Paulina; Waldor, Matthew K.; Chess, Andrew; Schadt, Eric E.; Fang, Gang

doi:10.1038/ncomms8438

Cited by 84 publications

(81 citation statements)

References 71 publications

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“…A host of assays have been developed to detect specific modified nucleotides, including and especially 5mC and 6mA, which are widely deployed by prokaryotes and eukaryotes [2][3][4] . Techniques exist to detect a diverse group of epigenetic modifications through the observation of DNA Pol II kinetics leveraging Single Molecule Real-Time sequencing (SMRT-seq) platform 5,6 . In particular, pioneering work 7,8 demonstrated the capacity to identify DNA methylation marks via the comparison of native versus amplified DNA through supervised machine learning.…”

Section: Introductionmentioning

confidence: 99%

De novoIdentification of DNA Modifications Enabled by Genome-Guided Nanopore Signal Processing

Stoiber

Quick

Egan

et al. 2016

Preprint

265

378

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Advances in nanopore sequencing technology have enabled investigation of the full catalogue of covalent DNA modifications. We present the first algorithm for the identification of modified nucleotides without the need for prior training data along with the open source software implementation, nanoraw. Nanoraw accurately assigns contiguous raw nanopore signal to genomic positions, enabling novel data visualization, and increasing power and accuracy for the discovery of covalently modified bases in native DNA. Ground truth case studies utilizing synthetically methylated DNA show the capacity to identify three distinct methylation marks, 4mC, 5mC, and 6mA, in seven distinct sequence contexts without any changes to the algorithm. We demonstrate quantitative reproducibility simultaneously identifying 5mC and 6mA in native E. coli across biological replicates processed in different labs. Finally we propose a pipeline for the comprehensive discovery of DNA modifications in any genome without a priori knowledge of their chemical identities.

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Section: Introductionmentioning

confidence: 99%

De novoIdentification of DNA Modifications Enabled by Genome-Guided Nanopore Signal Processing

Stoiber

Quick

Egan

et al. 2016

Preprint

265

378

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“…One of the central questions of cuprate physics is the role of competing electronic ordering tendencies in determining both normal-state and superconducting properties. Recently, charge-stripe order has emerged as a ubiquitous phenomenon in cuprates1234567 and charge stripes have been proposed to play a role in almost all of the important features of the cuprate phase diagram: the mysterious pseudogap phase89, Fermi surface reconstruction10 and even superconducting pairing11. However, firm experimental evidence for these claims is difficult to obtain, because charge stripes are sensitive to disorder, which is a prominent feature of all cuprate materials12.…”

mentioning

confidence: 99%

Unconventional charge order in a co-doped high-Tc superconductor

et al. 2016

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Charge-stripe order has recently been established as an important aspect of cuprate high-Tc superconductors. However, owing to the complex interplay between competing phases and the influence of disorder, it is unclear how it emerges from the parent high-temperature state. Here we report on the discovery of an unconventional ordered phase between charge-stripe order and (pseudogapped) metal in the cuprate La1.8−xEu0.2SrxCuO4. We use three complementary experiments—nuclear quadrupole resonance, nonlinear conductivity and specific heat—to demonstrate that the order appears through a sharp phase transition and exists in a dome-shaped region of the phase diagram. Our results imply that the new phase is a state, which preserves translational symmetry: a charge nematic. We thus resolve the process of charge-stripe development in cuprates, show that this nematic phase is distinct from high-temperature pseudogap and establish a link with other strongly correlated electronic materials with prominent nematic order.

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“…The broadness of signal perturbation suggests that methylation induces current differences across multiple flanking bases, essentially due to DNA methylation disturbing the ionic current of multiple consecutive events while ratcheting through the nanopore 3,4 . It is worth noting that this broadness contrasts with the deviations of kinetic DNA polymerase confined to a single base for 4mC and 6mA in SMRT sequencing 2,[20][21][22][23] .…”

Section: Heterogeneous Signal Variation Induced By Dna Methylation Inmentioning

confidence: 89%

Discovering and exploiting multiple types of DNA methylation from individual bacteria and microbiome using nanopore sequencing

Tourancheau

Mead

Zhang

et al. 2020

Preprint

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Nanopore sequencing provides a great opportunity for direct detection of chemical DNA modification. However, existing computational methods were either trained for detecting a specific form of DNA modification from one, or a few, specific sequence contexts (e.g. 5-methylcytosine from CpG dinucleotides) or for allowing de novo detection without effectively differentiating between different forms of DNA modifications. As a result, none of these methods supports de novo, systematic study of unknown bacterial methylomes. In this work, by examining three types of DNA methylation in a large diversity of sequence contexts, we observed that nanopore sequencing signal displays complex heterogeneity across methylation events of the same type. To capture this complexity and enable nanopore sequencing for broadly applicable methylation discovery, we generated a training dataset from an assortment of bacterial species and developed a novel method that couples the identification and fine mapping of the three forms of DNA methylation into a multi-label classification design. We evaluated the method and then applied it to individual bacteria and mouse gut microbiome for reliable methylation discovery. In addition, we demonstrated in the microbiome analysis the use of DNA methylation for binning metagenomic contigs, associating mobile genetic elements with their host genomes, and for the first time, identifying misassembled metagenomic contigs. This novel method has broad utility for discovering different forms of DNA methylation from bacteria, assisting functional studies of epigenetic regulation in bacteria, and exploiting bacterial epigenomes for more effective metagenomic analyses.

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Single molecule-level detection and long read-based phasing of epigenetic variations in bacterial methylomes

Cited by 84 publications

References 71 publications

De novoIdentification of DNA Modifications Enabled by Genome-Guided Nanopore Signal Processing

De novoIdentification of DNA Modifications Enabled by Genome-Guided Nanopore Signal Processing

Unconventional charge order in a co-doped high-Tc superconductor

Discovering and exploiting multiple types of DNA methylation from individual bacteria and microbiome using nanopore sequencing

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