Enabling NGS-based CRISPR QC applications with robust, high-performance DNA library preparation workflows
Jason Liu, PhD
Wednesday, December 5, 2018
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Chapters
Introduction
Agenda
How does CRISPR/Cas9 gene editing work?
Application of KAPA HyperPrep workflow to CRISPR/Cas9 gene editing verification
CRISPR/Cas9 on-target gene editing verification
High-throughput method overview
Input sample variability
Successful library construction
Quality assessment, sequencing set-up and results
Summary
CIRCLE-seq: a highly sensitive in vitro screen for genome-wide CRISPR–Cas9 nuclease off-targets
Comparing CRISPR–Cas9 off-target identification assays
CIRCLE-seq: in vitro genome-wide off-target identification method
High sensitivity requires high-fidelity enzymes to minimize artifacts
CIRCLE-seq is more sensitive than Digenome-seq
CIRCLE-seq is more sensitive than GUIDE-seq
Subset of off-target sites identified by CIRCLE-seq are verified to be mutated in human cells as well
Summary
Final Summary
Acknowledgements
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