Long-read single-molecule maps of the functional methylome

Sharim, Hila; Grunwald, Assaf; Gabrieli, Tslil; Michaeli, Yael; Margalit, Sapir; Torchinsky, Dmitry; Arielly, Rani; Nifker, Gil; Juhasz, Matyas; Gularek, Felix; Almalvez, Miguel; Dufault, Brandon; Chandra, Sreetama Sen; Liu, Alexander; Bhattacharya, Surajit; Chen, Yi Wen; Vilain, Éric; Wagner, Kathryn R.; Pevsner, Jonathan; Reifenberger, Jeff G.; Lam, Ernest T.; Hastie, Alex; Cao, Han; Barseghyan, Hayk; Weinhold, Elmar G.; Ebenstein, Yuval

doi:10.1101/gr.240739.118

Cited by 52 publications

(67 citation statements)

References 67 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…By combining the damage assay with optical DNA mapping it is possible to identify where along the genome each damage site is located [ 64 ]. This could be combined with commercial systems for optical DNA mapping to investigate genome-wide levels of DNA damage, similar to what was recently demonstrated for the epigenetic markers 5-methylcytosine [ 65 ] and 5-hydroxymethylcytosine [ 66 ].…”

Section: Discussionmentioning

confidence: 80%

Quantifying DNA damage induced by ionizing radiation and hyperthermia using single DNA molecule imaging

Singh

Johansson

Torchinsky

et al. 2020

Translational Oncology

Self Cite

View full text Add to dashboard Cite

Ionizing radiation (IR) is a common mode of cancer therapy, where DNA damage is the major reason of cell death. Here, we use an assay based on fluorescence imaging of single damaged DNA molecules isolated from radiated lymphocytes, to quantify IR induced DNA damage. The assay uses a cocktail of DNA-repair enzymes that recognizes and excises DNA lesions and then a polymerase and a ligase incorporate fluorescent nucleotides at the damage sites, resulting in a fluorescent “spot” at each site. The individual fluorescent spots can then be counted along single stretched DNA molecules and the global level of DNA damage can be quantified. Our results demonstrate that inclusion of the human apurinic/apyrimidinic endonuclease 1 (APE1) in the enzyme cocktail increases the sensitivity of the assay for detection of IR induced damage significantly. This optimized assay also allowed detection of a cooperative increase in DNA damage when IR was combined with mild hyperthermia, which is sometimes used as an adjuvant in IR therapy. Finally, we discuss how the method may be used to identify patients that are sensitive to IR and other types of DNA damaging agents.

show abstract

Section: Discussionmentioning

confidence: 80%

Quantifying DNA damage induced by ionizing radiation and hyperthermia using single DNA molecule imaging

Singh

Johansson

Torchinsky

et al. 2020

Translational Oncology

Self Cite

View full text Add to dashboard Cite

show abstract

“…Furthermore, we would prefer one combined report for the SV and CNV outputs coming from two different algorithms, and think that data would be best visualized by a composite ideogram style, such as described in 38 , and by ‘genome-wide CNV profiles’ similar to the CNV-microarrays that are now available within the most recent update of the Bionano software (Supplementary Figure 4). Finally, other very intriguing new developments include the potential for ‘phased’ detection of SVs and methylation, 39 which can offer additional value for cancer research and diagnostics. 40…”

Section: Discussionmentioning

confidence: 99%

Next generation cytogenetics: comprehensive assessment of 48 leukemia genomes by genome imaging

Neveling

Mantere

Vermeulen

et al. 2020

Preprint

View full text Add to dashboard Cite

Somatic structural variants are important for cancer development and progression. In a diagnostic set-up, especially for hematological malignancies, the comprehensive analysis of all cytogenetic aberrations in a given sample still requires a combination of techniques, such as karyotyping, fluorescence in situ hybridization and CNV-microarrays. We hypothesize that the combination of these classical approaches could be replaced by high-resolution genome imaging.Bone marrow aspirates or blood samples derived from 48 patients with leukemia, who received a clinical diagnoses of different types of hematological malignancies, were processed for genome imaging with the Bionano Genomics Saphyr system. In all cases cytogenetic abnormalities had previously been identified using standard of care workflows. Based on these diagnostic results, the samples were divided into two categories: simple cases (<5 aberrations, n=37) and complex cases (≥5 aberrations or an unspecified marker chromosome, n=11). By imaging the labelled ultra-long gDNA molecules (average N50 >250kb), we generated on average ∼280-fold mapped genome coverage per sample. Chromosomal aberrations were called by Bionano Genomics Rare variant pipeline (RVP) specialized for the detections of somatic variants.Per sample, on average a total of 1,454 high confidence SVs were called, and on average 44 (range: 14-130) of those were rare i.e. not present in the population control database. Importantly, for the simple cases, all clinically reported aberrations with variant allele frequencies higher than 10% were detected by genome imaging. This held true for deletions, insertions, inversions, aneuploidies and translocations. The results for the complex cases were also largely concordant between the standard of care workflow and optical mapping, and in several cases, optical mapping revealed higher complexity than previously known. SV and CNV calls detected by optical mapping were more complete than any other previous single test and likely delivered the most accurate and complete underlying genomic architecture. Even complex chromothripsis structures were resolved. Finally, optical mapping also identified multiple novel events, including balanced translocations that lead to potential novel fusion-genes, opening the potential to discover new prognostic and diagnostic biomarkers.The full concordance with diagnostic standard assays for simple cases and the overall great concordance with (previously likely incompletely understood) complex cases demonstrates the potential to replace classical cytogenetic tests with genome imaging. In addition, this holds the potential to rapidly map new fusion genes and identify novel SVs and CNVs as novel potential leukemia drivers.

show abstract

“…In 2018, Gabrieli et al [135] looked for 5-hydromethylcytosine, a product of active DNA demethylation, on the Bionano platform. Later, Sharim et al [136] advanced methylation detection by using a synthetic analog to “trick” methylases into transferring a fluorophore instead of a methyl group. Hence, the protocol is compatible with the high-throughput Bionano platform with the merit of minimizing DNA damage, similar to the DLS method discussed above.…”

Section: Applications Of Optical Mappingmentioning

confidence: 99%

Advances in optical mapping for genomic research

Yuan

Chung

Chan

2020

Computational and Structural Biotechnology Journal

View full text Add to dashboard Cite

Recent advances in optical mapping have allowed the construction of improved genome assemblies with greater contiguity. Optical mapping also enables genome comparison and identification of large-scale structural variations. Association of these large-scale genomic features with biological functions is an important goal in plant and animal breeding and in medical research. Optical mapping has also been used in microbiology and still plays an important role in strain typing and epidemiological studies. Here, we review the development of optical mapping in recent decades to illustrate its importance in genomic research. We detail its applications and algorithms to show its specific advantages. Finally, we discuss the challenges required to facilitate the optimization of optical mapping and improve its future development and application.

show abstract

Long-read single-molecule maps of the functional methylome

Cited by 52 publications

References 67 publications

Quantifying DNA damage induced by ionizing radiation and hyperthermia using single DNA molecule imaging

Quantifying DNA damage induced by ionizing radiation and hyperthermia using single DNA molecule imaging

Next generation cytogenetics: comprehensive assessment of 48 leukemia genomes by genome imaging

Advances in optical mapping for genomic research

Contact Info

Product

Resources

About