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Journal article
Customized optical mapping by CRISPR–Cas9 mediated DNA labeling with multiple sgRNAs
Nucleic acids research, v 49(2), pp e8-e8
25 Jan 2021
PMID: 33231685
Featured in Collection : UN Sustainable Development Goals @ Drexel
Abstract
Abstract
Whole-genome mapping technologies have been developed as a complementary tool to provide scaffolds for genome assembly and structural variation analysis (1,2). We recently introduced a novel DNA labeling strategy based on a CRISPR–Cas9 genome editing system, which can target any 20bp sequences. The labeling strategy is specifically useful in targeting repetitive sequences, and sequences not accessible to other labeling methods. In this report, we present customized mapping strategies that extend the applications of CRISPR–Cas9 DNA labeling. We first design a CRISPR–Cas9 labeling strategy to interrogate and differentiate the single allele differences in NGG protospacer adjacent motifs (PAM sequence). Combined with sequence motif labeling, we can pinpoint the single-base differences in highly conserved sequences. In the second strategy, we design mapping patterns across a genome by selecting sets of specific single-guide RNAs (sgRNAs) for labeling multiple loci of a genomic region or a whole genome. By developing and optimizing a single tube synthesis of multiple sgRNAs, we demonstrate the utility of CRISPR–Cas9 mapping with 162 sgRNAs targeting the 2Mb Haemophilus influenzae chromosome. These CRISPR–Cas9 mapping approaches could be particularly useful for applications in defining long-distance haplotypes and pinpointing the breakpoints in large structural variants in complex genomes and microbial mixtures.
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Details
- Title
- Customized optical mapping by CRISPR–Cas9 mediated DNA labeling with multiple sgRNAs
- Creators
- Heba Z Abid - Drexel UniversityEleanor Young - Drexel UniversityJennifer McCaffrey - Drexel UniversityKaitlin Raseley - Drexel UniversityDharma Varapula - Drexel UniversityHung-Yi Wang - Drexel UniversityDanielle Piazza - Drexel UniversityJoshua Mell - Drexel UniversityMing Xiao - Drexel University
- Publication Details
- Nucleic acids research, v 49(2), pp e8-e8
- Publisher
- Oxford University Press
- Resource Type
- Journal article
- Language
- English
- Academic Unit
- Biochemistry and Molecular Biology; Microbiology and Immunology; School of Biomedical Engineering, Science, and Health Systems
- Web of Science ID
- WOS:000626723800002
- Scopus ID
- 2-s2.0-85100358541
- Other Identifier
- 991019169705304721
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- Web of Science research areas
- Biochemistry & Molecular Biology