|
Status |
Public on Jan 07, 2019 |
Title |
CRISPR-based multimodal genetic screens in human iPSC-derived neurons |
Organism |
Homo sapiens |
Experiment type |
Expression profiling by high throughput sequencing
|
Summary |
CRISPR/Cas9-based functional genomics have transformed our ability to elucidate mammalian cell biology. Most previous CRISPR-based screens were implemented in cancer cell lines, rather than healthy, differentiated cells. Here, we describe a CRISPR interference (CRISPRi)-based platform for genetic screens in human neurons derived from induced pluripotent stem cells (iPSCs). We demonstrate robust and durable knockdown of endogenous genes in such neurons, and present results from three complementary genetic screens. A survival-based screen revealed neuron-specific essential genes and a small number of genes that improved neuronal survival upon knockdown. A screen with a single-cell transcriptomic readout uncovered several examples for genes knockdown of which had dramatically different cell-type specific consequences. A longitudinal imaging screen detected distinct consequences of gene knockdown on neuronal morphology. Our results highlight the potential of interrogating cell biology in iPSC-derived differentiated cell types and provide a platform for the systematic dissection of normal and disease states of neurons.
|
|
|
Overall design |
The Quant-seq experiments were performed using iPSC-derived neurons at Day 14, Day 21 and Day 28 of differentiation with 2 replicates for each sample. The CROP-seq experiments were performed using iPSCs and Day 7 neurons infected by a library of sgRNAs. Two lanes of 10X platform were used for each CROP-seq sample.
|
|
|
Contributor(s) |
Tian R, Ludwig C, Kampmann M |
Citation(s) |
31422865 |
Submission date |
Jan 06, 2019 |
Last update date |
Jun 19, 2020 |
Contact name |
Ruilin Tian |
E-mail(s) |
Ruilin.Tian@ucsf.edu
|
Organization name |
University of California, San Francisco
|
Department |
Institute for Neurodegenerative Diseases
|
Lab |
Martin Kampmann
|
Street address |
675 Nelson Rising Lane, 3rd Floor
|
City |
San Francisco |
State/province |
CA |
ZIP/Postal code |
94158 |
Country |
USA |
|
|
Platforms (2) |
GPL20301 |
Illumina HiSeq 4000 (Homo sapiens) |
GPL24676 |
Illumina NovaSeq 6000 (Homo sapiens) |
|
Samples (14)
|
|
Relations |
BioProject |
PRJNA513160 |
SRA |
SRP176013 |