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2023 ACMG Annual Clinical Genetics Meeting Digital ...
Resolving clinically relevant short read deficient ...
Resolving clinically relevant short read deficient homologous sequences using a novel CRISPR-CAS mediated targeted long read sequencing method
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This document describes a novel method using the CRISPR-Cas9 system for targeted long read sequencing. The method involves the use of CRISPR-Cas9 cleavage to enrich for specific regions of interest in the genome. The study first demonstrates the effectiveness of CRISPR-Cas9 cleavage for depleting long cDNA fragments in an E. coli isolate. The results show a 99.2% depletion rate of targeted coding ribosomal and mitochondrial RNA transcripts using this method. The study also shows that CRISPR-Cas9 cleavage can be applied to fragments bound to beads, providing the ability to highly enrich for regions of interest.<br /><br />The study then extends the use of CRISPR-Cas9 cleavage to human PBMC samples. The method effectively depletes targeted coding ribosomal and mitochondrial transcripts and exhibits low bias between control and depleted samples. The study concludes that combining CRISPR-Cas9 cleavage and enrichment with long read sequencing can provide a novel approach for enriching clinically relevant genes with complex homology while generating full-length information across the gene.<br /><br />Future work will focus on combining these methods into a single workflow that can be used with short or long read sequencing systems. The document includes various figures and diagrams illustrating the workflow and results of the study.<br /><br />Overall, this study demonstrates the potential of the CRISPR-Cas9 system for targeted long read sequencing and highlights its effectiveness in depleting specific gene transcripts.
Asset Subtitle
Presenting Author - Madhuri Hegde, PhD, FACMG; Co-Author - Fen Guo, PhD, FACMG, FCCMG; Co-Author - Jorune Balciuniene, PhD, FACMG, ErCLG; Co-Author - S Ranganathan, PhD; Co-Author - J Armstrong, PhD; Co-Author - Keith Brown, PhD; Co-Author - A Siddique, PhD; Co-Author - Zeqiang Ma, PhD;
Meta Tag
Bioinformatics
Clinical Applications of Molecular Cytogenetics
Clinical Cytogenetics
Comparative Mapping
Cytogenetics
Databases
Education
Gene Localization
Gene Mapping
Genetic Testing
Genome sequencing
Genomic Methodologies
Genomic Structure
Methodology
NextGen Sequencing
Sequencing
Structure/Function
Co-Author
Fen Guo, PhD, FACMG, FCCMG
Co-Author
Jorune Balciuniene, PhD, FACMG, ErCLG
Co-Author
S Ranganathan, PhD
Co-Author
J Armstrong, PhD
Co-Author
Keith Brown, PhD
Co-Author
A Siddique, PhD
Co-Author
Zeqiang Ma, PhD
Presenting Author
Madhuri Hegde, PhD, FACMG
Keywords
CRISPR-Cas9 system
targeted long read sequencing
CRISPR-Cas9 cleavage
genome enrichment
E. coli isolate
depletion rate
coding ribosomal RNA
mitochondrial RNA
bead-bound fragments
human PBMC samples
© 2024 American College of Medical Genetics and Genomics. All rights reserved.
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