Figure 5.

Efficient gene editing of primary cells by a.Gag.MS2.CRISPR particles

(A) Highly efficient knockout of endogenous TP53 in primary human NuFF cells. NuFF cells were transduced with serial dilutions of a.Gag.MS2.CRISPR.TP53 particles reflecting the indicated particle doses expressed as SpCas9.TS mRNA copies per cell. The knockout efficiencies were determined by TIDE analysis. (B) TP53 knockout cells showed enhanced proliferation compared with non-treated mock cells. Cells from (A) were counted and seeded at a density of 1 × 10⁴ per well. Seven to eight days later, the respective total number of cells was determined. (C and D) TP53 knockout rates in primary human hepatocytes (PHHs) and human umbilical cord blood-derived CD34⁺ hematopoietic stem and progenitor cells (hCD34⁺ HSPCs). (E) Efficient knockout of endogenous Trp53 in murine embryonic fibroblasts (MEFs). CF1 or C3H MEFs were transduced with 4,800 SpCas9.TS mRNA copies/cell on 2 consecutive days. The data points in respective graphs reflect biological replicates with independently prepared supernatants (n = 2–4).