|
| Status |
Public on Nov 18, 2019 |
| Title |
Titrating gene expression with allelic series of CRISPR guide RNAs |
| Organism |
Homo sapiens |
| Experiment type |
Expression profiling by high throughput sequencing
|
| Summary |
Cellular phenotypes arise from the degrees to which different genes are expressed, yet the ability to monitor these phenotypes is limited by a lack of tools to precisely control gene expression. We describe the development of allelic series of systematically compromised sgRNAs to titrate expression of human genes with CRISPR interference. Using large-scale measurements of compromised sgRNA activities, we both identify empirically validated intermediate-activity sgRNAs and derive the factors governing sgRNA activity using deep learning, enabling construction of a compact sgRNA library to titrate expression of ~2,400 essential genes and a genome-wide in silico library. Staging cells along a continuum of essential gene expression levels using sgRNA series combined with rich single-cell RNA-seq readout reveals expression threshold-specific responses and gene-specific expression-to-phenotype relationships. Our work provides a general tool to control gene expression with applications ranging from tuning of biochemical pathways to identification of suppressors for diseases of dysregulated gene expression.
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| |
|
| Overall design |
Pooled CRISPR screening experiment with intermediate-activity sgRNAs conducted via Perturb-seq
|
| |
|
| Contributor(s) |
Jost M, Santos DA, Saunders RA, Weissman JS |
| Citation(s) |
31932729 |
| NIH grant(s) |
| Grant ID |
Grant title |
Affiliation |
Name |
| U01 CA168370 |
Bay Area Cancer Target Discovery and Development Network |
University of California San Francisco |
JONATHAN S. WEISSMAN |
| U01 CA217882 |
The Cancer Target Discovery and Development Network at UCSF |
University of California San Francisco |
JONATHAN S. WEISSMAN |
| F32 GM116331 |
Systematic Genome-Wide Characterization of Iron Homeostasis |
University of California San Francisco |
Marco Jost |
| K99 GM130964 |
Deciphering the logic of glycolipid signaling at the host-microbiome interface |
University of California San Francisco |
Marco Jost |
| RM1 HG009490 |
Center for Genome Editing and Recording |
UNIVERSITY OF CALIFORNIA BERKELEY |
JENNIFER A DOUDNA |
|
| |
| Submission date |
Jun 03, 2019 |
| Last update date |
Jan 15, 2020 |
| Contact name |
Marco Jost |
| E-mail(s) |
marco_jost@hms.harvard.edu
|
| Organization name |
Harvard Medical School
|
| Street address |
77 Ave Louis Pasteur, NRB848
|
| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02115 |
| Country |
USA |
| |
|
| Platforms (1) |
| GPL20301 |
Illumina HiSeq 4000 (Homo sapiens) |
|
| Samples (6)
|
| GSM3842207 |
Intermediate-activity sgRNA perturb-seq experiment (gemgroup1) |
| GSM3842208 |
Intermediate-activity sgRNA perturb-seq experiment (gemgroup2) |
| GSM3842209 |
Intermediate-activity sgRNA perturb-seq experiment (gemgroup3) |
| GSM3842210 |
Intermediate-activity sgRNA perturb-seq experiment sgRNA barcode reads (gemgroup1) |
| GSM3842211 |
Intermediate-activity sgRNA perturb-seq experiment sgRNA barcode reads (gemgroup2) |
| GSM3842212 |
Intermediate-activity sgRNA perturb-seq experiment sgRNA barcode reads (gemgroup3) |
|
| Relations |
| BioProject |
PRJNA545894 |
| SRA |
SRP200150 |
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