Related Experiment Video
Updated: Apr 12, 2026
14:06
Detection of Rare Genomic Variants from Pooled Sequencing Using SPLINTER
Published on: June 23, 2012
15.1K
High-throughput screening of human genetic variants by pooled prime editing
Michael Herger1, Christina M Kajba1, Megan Buckley1
1The Genome Function Laboratory, The Francis Crick Institute, London NW1 1AT, UK.
Cell Genomics
|March 22, 2025
Summary
We developed a novel pooled prime editing platform to efficiently screen thousands of genetic variants in their native cellular environment. This method accurately identifies loss-of-function variants, advancing human genetic disease research.
Area of Science:
- Genetics
- Molecular Biology
- Bioinformatics
Background:
- Multiplexed assays of variant effect (MAVEs) are crucial for assessing human genetic variants.
- Existing MAVEs face limitations due to exogenous expression or genome editing constraints.
Purpose of the Study:
- To introduce and validate a pooled prime editing (PE) platform for scalable, endogenous variant effect assays.
- To improve PE efficiency and establish robust screening methods for loss-of-function (LoF) variants.
Main Methods:
- Optimized prime editing guide RNA (pegRNA) design and co-selection strategies in HAP1 cells.
- Negative selection screening of thousands of pegRNAs targeting SMARCB1 and MLH1 genes.
- Assessed single nucleotide variants (SNVs) and ClinVar non-coding variants across large genomic regions.
Main Results:
- Demonstrated efficient installation and negative selection of loss-of-function variants in SMARCB1.
- Successfully screened a large number of SNVs and non-coding variants in MLH1 using 6-thioguanine selection.
- The platform showed high accuracy in discriminating pathogenic variants.
Conclusions:
- The pooled prime editing platform enables scalable, endogenous assessment of genetic variant effects.
- This technology is valuable for identifying novel pathogenic variants associated with human phenotypes across extensive genomic regions.
