Frequent sgRNA-barcode recombination in single-cell perturbation assays

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

Research output: Contribution to journalArticle

11 Citations (Scopus)

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)
Article numbere0198635
JournalPLoS One
Volume13
Issue number6
DOIs
StatePublished - Jun 1 2018

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barcoding
Genetic Recombination
Assays
Clustered Regularly Interspaced Short Palindromic Repeats
assays
Virus Assembly
Signal to noise ratio
Packaging
Genes
Throughput
Signal-To-Noise Ratio
Proxy
cells
Libraries
Genome
packaging
genome

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)

Cite this

Frequent sgRNA-barcode recombination in single-cell perturbation assays. / Xie, Shiqi; Cooley, Anne; Armendariz, Daniel; Zhou, Pei; Hon, Gary C.

In: PLoS One, Vol. 13, No. 6, e0198635, 01.06.2018.

Research output: Contribution to journalArticle

Xie, Shiqi ; Cooley, Anne ; Armendariz, Daniel ; Zhou, Pei ; Hon, Gary C. / Frequent sgRNA-barcode recombination in single-cell perturbation assays. In: PLoS One. 2018 ; Vol. 13, No. 6.
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