Frequent sgRNA-barcode Recombination in Single-cell Perturbation Assays

Shiqi Xie, Anne Cooley, Daniel Armendarez, Pei Zhou, Gary C. Hon

Research output: Contribution to journalArticlepeer-review

Abstract

Simultaneously detecting CRISPR-based perturbations and induced transcriptional changes in the same cell is a powerful approach to unraveling genome function. Several lentiviral approaches have been developed, some of which rely on the detection of distally located genetic barcodes as an indirect proxy of sgRNA identity. Since barcodes are often several kilobases from their corresponding sgRNAs, viral recombination-mediated swapping of barcodes and sgRNAs is feasible. Using a self-circularization-based sgRNA-barcode library preparation protocol, we estimate the recombination rate to be ~50% and we trace this phenomenon to the pooled viral packaging step. Recombination is random, and decreases the signal-to-noise ratio of the assay. Our results suggest that alternative approaches can increase the throughput and sensitivity of single-cell perturbation assays.

Original languageEnglish (US)
JournalUnknown Journal
DOIs
StatePublished - Jan 29 2018

Keywords

  • Barcode Shuffling
  • CRISP-seq
  • CRISPR
  • CROP-sq
  • Mosaic-seq
  • Perturb-seq
  • scRNA-seq

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)
  • Immunology and Microbiology(all)
  • Neuroscience(all)
  • Pharmacology, Toxicology and Pharmaceutics(all)

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