Single-molecule optical genome mapping in nanochannels: multidisciplinarity at the nanoscale

Abstract

The human genome contains multiple layers of information that extend beyond the genetic sequence. In fact, identical genetics do not necessarily yield identical phenotypes as evident for the case of two different cell types in the human body. The great variation in structure and function displayed by cells with identical genetic background is attributed to additional genomic information content. This includes large-scale genetic aberrations, as well as diverse epigenetic patterns that are crucial for regulating specific cell functions. These genetic and epigenetic patterns operate in concert in order to maintain specific cellular functions in health and disease. Single-molecule optical genome mapping is a high-throughput genome analysis method that is based on imaging long chromosomal fragments stretched in nanochannel arrays. The access to long DNA molecules coupled with fluorescent tagging of various genomic information presents a unique opportunity to study genetic and epigenetic patterns in the genome at a single-molecule level over large genomic distances. Optical mapping entwines synergistically chemical, physical, and computational advancements, to uncover invaluable biological insights, inaccessible by sequencing technologies. Here we describe the method’s basic principles of operation, and review the various available mechanisms to fluorescently tag genomic information. We present some of the recent biological and clinical impact enabled by optical mapping and present recent approaches for increasing the method’s resolution and accuracy. Finally, we discuss how multiple layers of genomic information may be mapped simultaneously on the same DNA molecule, thus paving the way for characterizing multiple genomic observables on individual DNA molecules.

Long reads capture simultaneous enhancer–promoter methylation status for cell-type deconvolution

“…Label coordinates were extended to 1 kb to account for the optical measurement resolution, limiting localization accuracy to ∼1000 bp ( Wang et al , 2012 ). More details regarding ROM and optical mapping can be found in ( Jeffet et al , 2021 ; Sharim et al , 2019 ).…”

Evaluation of optical genome mapping for detecting chromosomal translocation in clinical cytogenetics

“…Bionano optical genome mapping (OGM) is a recently developed technique can directly image labeled DNA molecules. Endonucleases are used in Bionano OGM instruments to recognize DNA sequences and cut single strands, so as to simultaneously label fluorescence and subsequently straightening, linearizing, and unfolding each single DNA molecule by very fine capillary electrophoresis for high‐resolution fluorescence imaging of ultra‐long single molecules (Jeffet et al, 2021 ; Schwartz et al, 1993 ). The structure of chromosomes can be analyzed according to the order of fluorescence signals on DNA molecules.…”

Engineering Infrequent DNA Nicking Endonuclease by Fusion of a BamHI Cleavage-Deficient Mutant and a DNA Nicking Domain

“…Sequence-specific DNA NEases have been used in DNA optical mapping by nicking, nick translation of the nicked sites with fluorescently labeled dNTP, and religation of the nicked strand (Zhang et al, 2010;Jeffet et al, 2021). Large chromosome rearrangement (insertion, deletion, or translocation) of cancer cells could be "visualized" under high-resolution fluorescent microscope when the abnormal DNA nicking pattern images are compared with the WT (English et al, 2015;Jeffet et al, 2021) (Bionano Genomics, Irys system).…”

“…Sequence-specific DNA NEases have been used in DNA optical mapping by nicking, nick translation of the nicked sites with fluorescently labeled dNTP, and religation of the nicked strand (Zhang et al, 2010;Jeffet et al, 2021). Large chromosome rearrangement (insertion, deletion, or translocation) of cancer cells could be "visualized" under high-resolution fluorescent microscope when the abnormal DNA nicking pattern images are compared with the WT (English et al, 2015;Jeffet et al, 2021) (Bionano Genomics, Irys system). NEases are also used in isothermal DNA amplifications, such as strand-displacement DNA amplification (SDA) (Walker et al, 1994), EXPAR (Qian et al, 2012), nicking enzyme assisted amplification (NEAA) (Van Ness et al, 2003;Qian et al, 2019), Nt.CviPII-assisted random whole genome amplification (Chan et al, 2004), and in NicE-seq to study open chromatin profiling (Ponnaluri et al, 2017).…”