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. 2021 Dec 29;221(2):e202104046. doi: 10.1083/jcb.202104046

Figure S1.

Figure S1.

Genetic engineering of HL-60 cells. (A) Strategy for endogenous tagging of WASP with split mNG2 using a Cas9–sgRNA RNP complex and ssDNA donor. (B) MiSeq results from A reveal high cutting efficiency in HL-60s but poor homology-directed repair (HDR). Analysis of paired end reads was done with CRISPResso2 (Clement et al., 2019). (C) Sequencing of WASP KI cells confirms correct insertion of mNG211 tag. (D) Strategy for endogenous tagging of clathrin LCa with full-length TagRFP-T using a Cas9–sgRNA RNP complex and plasmid donor. (E) Sequencing of an isolated homozygous CLTA KI clone shows correct repair at both ends of the cut site. (F) Sequence validation of the two clonal WASP KO lines assayed. Both lines have deletions that lead to a frame shift, nonsense, and termination following the end of exon 1. (G) Western blot confirms that WASP is absent in both assayed clonal lines. NHEJ, nonhomologous end joining.