2023 ACMG Annual Clinical Genetics Meeting Digital Edition: Poster Gallery-Resolving clinically relevant short read deficient homologous sequences using a novel CRISPR-CAS mediated targeted long read sequencing method

Resolving clinically relevant short read deficient homologous sequences using a novel CRISPR-CAS mediated targeted long read sequencing method

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Pdf Summary

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

Structure/Function

Sequencing

NextGen Sequencing

Methodology

Genomic Structure

Genomic Methodologies

Genome sequencing

Genetic Testing

Gene Mapping

Gene Localization

Education

Databases

Cytogenetics

Comparative Mapping

Clinical Cytogenetics

Clinical Applications of Molecular Cytogenetics

Bioinformatics

Co-Author Zeqiang Ma, PhD

Co-Author A Siddique, PhD

Co-Author Keith Brown, PhD

Co-Author J Armstrong, PhD

Co-Author S Ranganathan, PhD

Co-Author Jorune Balciuniene, PhD, FACMG, ErCLG

Co-Author Fen Guo, PhD, FACMG, FCCMG

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