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. Author manuscript; available in PMC: 2019 Jun 4.
Published in final edited form as: Nat Protoc. 2018 Jan 11;13(1):307–323. doi: 10.1038/nprot.2017.131

Figure 3. Assessing Cas9 Activity Using an sgRNA Targeting SAG1.

Figure 3.

(a) RH/Cas9 and RH are transfected with pU6-SAG1 or mock transfected, then cultured for 3 days. Pyrimethamine is added after 24 hours to select for transfected parasites (or not, in the case of mock transfected parasites). (b) Immunofluorescence assays (IFAs) performed after 3 days reveal loss of SAG1. Active Cas9 is expected to result in loss of SAG1 in 90–97% of parasite vacuoles. See steps 1–25 for a detailed procedure.