Journal: Nature Communications

Article Title: Evaluation of Cas13d as a tool for genetic interaction mapping

doi: 10.1038/s41467-025-56747-4

Figure Lengend Snippet: A Schematic of the different CRISPR systems, targeting DNA (Cas9 and Cas12a) or RNA (Cas13d). B Histograms of cells perturbed with Cas9, Cas12a, and Cas13d targeting CD46 with protein levels measured via flow cytometry at four different time points post-transduction of the respective CRISPR system. Light gray: Unstained untransduced cells. Red: Perturbed cells. Dark gray: Stained non-target control cells. C Gene perturbation kinetics of Cas9, Cas12a, and Cas13d targeting CD46, CD47, CD63, and CD71 over 10 days. Values represent the mean of biological replicates; error bars, SD ( n = 3). D Distribution of cell populations expressing combinations of CD46, CD47 and NTC gRNAs as indicated. Cell surface marker levels were quantified via flow cytometry at 10 days post-transduction. E Quantification of unperturbed, single-perturbed, and double-perturbed sub-populations from cells expressing gRNAs against CD46 and CD47 at 10 days post-transduction. Values represent the mean of biological replicates; error bars, SD ( n = 3). Source data are provided as a Source Data file.

Article Snippet: The mCD4 gene was amplified from the S. aureus Cas9 nuclease vector (Addgene #105998) using Phusion Flash High-Fidelity PCR Master Mix (ThermoFisher Scientific, F548L) according to the manufacturer’s protocol.

Techniques: CRISPR, Flow Cytometry, Transduction, Staining, Control, Expressing, Marker