Long-read single-molecule maps of the functional methylome.

We report on the development of a methylation analysis workflow for optical detection of fluorescent methylation profiles along chromosomal DNA molecules. In combination with Bionano Genomics genome mapping technology, these profiles provide a hybrid genetic/epigenetic genome-wide map composed of DNA molecules spanning hundreds of kilobase pairs. The method provides kilobase pair-scale genomic methylation patterns comparable to whole-genome bisulfite sequencing (WGBS) along genes and regulatory elements. These long single-molecule reads allow for methylation variation calling and analysis of large structural aberrations such as pathogenic macrosatellite arrays not accessible to single-cell second-generation sequencing. The method is applied here to study facioscapulohumeral muscular dystrophy (FSHD), simultaneously recording the haplotype, copy number, and methylation status of the disease-associated, highly repetitive locus on Chromosome 4q.

Epigenetic Optical Mapping of 5-Hydroxymethylcytosine in Nanochannel Arrays.

The epigenetic mark 5-hydroxymethylcytosine (5-hmC) is a distinct product of active DNA demethylation that is linked to gene regulation, development, and disease. In particular, 5-hmC levels dramatically decline in many cancers, potentially serving as an epigenetic biomarker. The noise associated with next-generation 5-hmC sequencing hinders reliable analysis of low 5-hmC containing tissues such as blood and malignant tumors. Additionally, genome-wide 5-hmC profiles generated by short-read sequencing are limited in providing long-range epigenetic information relevant to highly variable genomic regions, such as the 3.7 Mbp disease-related Human Leukocyte Antigen (HLA) region. We present a long-read, highly sensitive single-molecule mapping technology that generates hybrid genetic/epigenetic profiles of native chromosomal DNA. The genome-wide distribution of 5-hmC in human peripheral blood cells correlates well with 5-hmC DNA immunoprecipitation (hMeDIP) sequencing. However, the long single-molecule read-length of 100 kbp to 1 Mbp produces 5-hmC profiles across variable genomic regions that failed to show up in the sequencing data. In addition, optical 5-hmC mapping shows a strong correlation between the 5-hmC density in gene bodies and the corresponding level of gene expression. The single-molecule concept provides information on the distribution and coexistence of 5-hmC signals at multiple genomic loci on the same genomic DNA molecule, revealing long-range correlations and cell-to-cell epigenetic variation.

Irys Extract.

Summary: Irys Extract is a software tool for generating genomic information from data collected by the BioNano Genomics Irys platform. The tool allows the user easy access to the raw data in the form of cropped images and genetically aligned intensity profiles. The latter are also made compatible with the BED format for using with popular genomic browsers such as the UCSC Genome Browser. Availability and implementation: Irys Extract has been developed in Matlab R2015a, it was tested to work with IrysView 2.4.0.15879 and AutoDetect 2.1.4.9159, and it currently runs under Microsoft Windows operating systems (7-10). Irys Extract can be downloaded alongside its manual and a demo dataset at http://www.nanobiophotonix.com and https://sites.google.com/site/raniarielly/. Contact: uv@post.tau.ac.il. Supplementary information: Supplementary data are available at Bioinformatics online.

Real-time detection of redox events in molecular junctions.

Redox molecular junctions are promising systems for nanoelectronics applications, and yet they are still only marginally understood. The study of these systems has so far been conducted in solution, utilizing "electrolyte gating" to control their redox states and, as a result, their steady-state transistor-like conductance behavior. Here we explore redox junctions under vacuum at 77 K, and report real time detection of redox events in junctions of the type Au-6-thiohexanethiolferrocene-Au. Redox events are revealed as a two-level fluctuating signal in current-time traces with potential-dependent amplitude and frequency. Using a theoretical model for signals with a telegraph-like noise, the current-time traces are analyzed to extract the various molecular parameters which define the dynamics of the system. The presented method, which can be applied to other types of redox molecules, offers a new approach to study the unexplored territory of molecular dynamics in molecular junctions.

Accurate determination of plasmonic fields in molecular junctions by current rectification at optical frequencies.

Current rectification, i.e., induction of dc current by oscillating electromagnetic fields, is demonstrated in molecular junctions at an optical frequency. The magnitude of rectification is used to accurately determine the effective oscillating potentials in the junctions induced by the irradiating laser. Since the gap size of the junctions used in this study is precisely determined by the length of the embedded molecules, the oscillating potential can be used to calculate the plasmonic enhancement of the electromagnetic field in the junctions. With a set of junctions based on alkyl thiolated molecules with identical HOMO-LUMO gap and different lengths, an exponential dependence of the plasmonic field enhancement on gap size is observed.