CRISPR-UMI Step by Step: A protocol for robust CRISPR screening
CRISPR-UMI extends the existing repertoire of CRSIPR-screening methods. It circumvents cell heterogeneity, a consequence of Cas9 genome editing, by scoring single cell derived clones individually. The strength of this new CRISPR screening method is its robustness towards clonal heterogeneity and clonal outliers and is therefore expected to be most useful in challenging biological screens with strong bottlenecks and clonal effects such as organoid or in-vivo screens.
This step-by-step protocol is an addition to the publication CRISPR-UMI: Single cell lineage tracing of pooled CRISPR/Cas9 screens doi: 10.1038/nmeth.4466. It contains a detailed description for pooled CRISPR screening using CRISPR-UMI. It especially highlights the steps which are critical and unique to the use of CRISPR-UMI. Those critical steps are library preparation at very high complexity of up to 100million unique plasmids, and data analysis were unique guide-UMI pairs are evaluated separately.
This is a list of supplementary files associated with this preprint. Click to download.
Main document 1 CRISPR-UMI Step by Step Main Document This is the main document for the CRISPR-UMI Step by Step protocol.
Zip_archive 2 Scripts 1-23 A zip archive. contains all Scripts (perl, python, R, sh) mentioned in the Step by Step protocol.
Table 1 Primers_oligos table Excel file contains short Sequences Primers and oligo mentioned in the Step by Step protocol
Zip_arichive 3 Sequences 1-4 Vector maps and Sequence files in gbk format. mentioned in the Step by Step protocol.
Zip_archive 1 Files 1-20 A zip archive. contains all Files (Data Files, Intermediate Files and results) mentioned in the Step by Step protocols
Posted 17 Oct, 2017
CRISPR-UMI Step by Step: A protocol for robust CRISPR screening
Posted 17 Oct, 2017
CRISPR-UMI extends the existing repertoire of CRSIPR-screening methods. It circumvents cell heterogeneity, a consequence of Cas9 genome editing, by scoring single cell derived clones individually. The strength of this new CRISPR screening method is its robustness towards clonal heterogeneity and clonal outliers and is therefore expected to be most useful in challenging biological screens with strong bottlenecks and clonal effects such as organoid or in-vivo screens.
This step-by-step protocol is an addition to the publication CRISPR-UMI: Single cell lineage tracing of pooled CRISPR/Cas9 screens doi: 10.1038/nmeth.4466. It contains a detailed description for pooled CRISPR screening using CRISPR-UMI. It especially highlights the steps which are critical and unique to the use of CRISPR-UMI. Those critical steps are library preparation at very high complexity of up to 100million unique plasmids, and data analysis were unique guide-UMI pairs are evaluated separately.
© Research Square 2021