PT  - JOURNAL ARTICLE
AU  - Britt Adamson
AU  - Thomas M. Norman
AU  - Marco Jost
AU  - Jonathan S. Weissman
TI  - Approaches to maximize sgRNA-barcode coupling in Perturb-seq screens
AID  - 10.1101/298349
DP  - 2018 Jan 01
TA  - bioRxiv
PG  - 298349
4099  - http://biorxiv.org/content/early/2018/04/11/298349.short
4100  - http://biorxiv.org/content/early/2018/04/11/298349.full
AB  - Perturb-seq is a platform for single-cell gene expression profiling of pooled CRISPR screens. Like many functional genomics platforms, Perturb-seq relies on lentiviral transduction to introduce perturbation libraries to cells. On this platform, these are barcoded sgRNA libraries. A critical consideration for performing Perturb-seq experiments is uncoupling of barcodes from linked sgRNA expression cassettes, which can occur during lentiviral transduction of co-packaged libraries due to reverse transcriptase-mediated template switching. This problem is common to lentiviral libraries designed with linked variable regions. Here, we demonstrate that recombination between Perturb-seq vectors scrambles linked variable regions separated by 2 kb. This predicts information loss in Perturb-seq screens performed with co-packaged libraries. We also demonstrate ways to address this problem and discuss best practices for single-cell screens with transcriptional readouts.