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Journal article
High-throughput single-molecule telomere characterization
Genome research, v 27(11), pp 1904-1915
01 Nov 2017
PMID: 29025896
Featured in Collection : UN Sustainable Development Goals @ Drexel
Abstract
We have developed a novel method that enables global subtelomere and haplotype-resolved analysis of telomere lengths at the single-molecule level. An in vitro CRISPR/Cas9 RNA-directed nickase system directs the specific labeling of human (TTAGGG) n DNA tracts in genomes that have also been barcoded using a separate nickase enzyme that recognizes a 7bp motif genome-wide. High-throughput imaging and analysis of large DNA single molecules from genomes labeled in this fashion using a nanochannel array system permits mapping through subtelomere repeat element (SRE) regions to unique chromosomal DNA while simultaneously measuring the (TTAGGG) n tract length at the end of each large telomere- terminal DNA segment. The methodology also permits subtelomere and haplotype-resolved analyses of SRE organization and variation, providing a window into the population dynamics and potential functions of these complex and structurally variant telomere-adjacent DNA regions. At its current stage of development, the assay can be used to identify and characterize telomere length distributions of 30-35 discrete telomeres simultaneously and accurately. The assay's utility is demonstrated using early versus late passage and senescent human diploid fibroblasts, documenting the anticipated telomere attrition on a global telomere-by-telomere basis as well as identifying subtelomere-specific biases for critically short telomeres. Similarly, we present the first global single-telomere-resolved analyses of two cancer cell lines.
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Details
- Title
- High-throughput single-molecule telomere characterization
- Creators
- Jennifer McCaffrey - Drexel UniversityEleanor Young - Drexel UniversityKaty Lassahn - Old Dominion UniversityJustin Sibert - Drexel UniversitySteven Pastor - Drexel UniversityHarold Riethman - Old Dominion UniversityMing Xiao - Drexel University
- Publication Details
- Genome research, v 27(11), pp 1904-1915
- Publisher
- Cold Spring Harbor Lab Press, Publications Dept
- Number of pages
- 12
- Grant note
- R01CA140652 / NATIONAL CANCER INSTITUTE; United States Department of Health & Human Services; National Institutes of Health (NIH) - USA; NIH National Cancer Institute (NCI) FV7[L;]9 R21HG007205; R21CA177395 / US National Institutes of Health (NIH); United States Department of Health & Human Services; National Institutes of Health (NIH) - USA Drexel's University Research Computing Facility R21HG007205 / NATIONAL HUMAN GENOME RESEARCH INSTITUTE; United States Department of Health & Human Services; National Institutes of Health (NIH) - USA; NIH National Human Genome Research Institute (NHGRI)
- Resource Type
- Journal article
- Language
- English
- Academic Unit
- School of Biomedical Engineering, Science, and Health Systems
- Web of Science ID
- WOS:000414165900011
- Scopus ID
- 2-s2.0-85045286605
- Other Identifier
- 991019168088804721
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- Collaboration types
- Domestic collaboration
- Web of Science research areas
- Biochemistry & Molecular Biology
- Biotechnology & Applied Microbiology
- Genetics & Heredity