A clonal derivative of SVG-A expressing Cas9 and gRNA m1 has reduced capacity to support JCV infection.

A. SVG-A cells were transfected with Cas9 or Cas9 plus gRNA m1 and stable clonal cell lines selected. Representative clones were selected and assayed for JCV infection (MOI = 1, 7 days post-infection): results from one clone with Cas9 alone and one clone with Cas9 plus gRNA m1 (c8) relative to parental SVG-A cells. Viral infection was assessed by Western blot for VP1 and agnoprotein with α-tubulin as a loading control. B. Viral loads in the culture supernatants from the experiment in Panel A were quantified using Q-PCR and are shown as copy number. C. SVG-ACas9 and SVG-ACas9m1c8 were assayed for Cas9 expression by Western blot with α-tubulin used as a loading control. D. TC620 cells were transfected with expression plasmid for FLAG-tagged Cas9 and immunocytochemistry performed with anti-FLAG antibody as described in Materials and Methods. Nuclei were labeled with 4',6-diamidino-2-phenylindole (DAPI).