GSE108699
On the design of CRISPR-based single cell molecular screens
We demonstrate that vector designs for such screens that rely on cis linkage of guides and distally located barcodes suffer from swapping of intended guide-barcode associations at rates approaching 50% due to template switching during lentivirus production, greatly reducing sensitivity. Overall design: Experiments were conducted with FACs followed by NGS to quantify the rate of template switching. single-cell RNA-seq experiments were performed on the 10X Genomics platform for mock and doxorubicin treated MCF10a cells that had been tranduced with a CRISPR KO library (targeting tumor suppressors) that allowed for readout of the KO via a linked distal barcode using both an arrayed lentivirus production strategy and a pooled lentivirus production strategy. single-cell RNA-seq was also performed on mock and doxorubicin cells tranduced with a CRISPR KO library targeting tumor suppressors using a CROP-seq vector as the backbone.