Fig. 2

The genome of CHSE-EC salmon cells is efficiently edited with lentivirus. a-d Efficient editing of EGFP in CHSE-EC Chinook salmon cell line by lentivirus. CHSE-EC cells were spinfected for 2 h at 1000 g with neat (Hi) and 1:10 dilution (Lo) of lentivirus supernatant and incubated at 22 °C. After 2 weeks of expansion, puromycin was added to the Lo group (0.25 μg / mL, for 1 week, Lo + Puro). Fluorescence was imaged by epifluorescence microscopy (a) and recorded flow cytometry (b) using CHSE wt and CHSE-EC (not transduced) as controls. Split histogram of fluorescence and corresponding proportion histogram of control cells (top and bottom) and CHES-EC transduced with high and low concentration (with and without puromycin treatment) lentivirus supernatant. Scale bar in a represents 20 μm. c Genome editing efficiency in CHSE-EC by Sanger sequencing of PCR amplified product of the target loci, estimated by ICE analysis. d Detail of indels frequency in EGFP edited Hi group (from panel c), estimated by ICE analysis. Purple dot denotes the unedited sequence (0 bp). e-f Efficient editing of RIG-I in CHSE-EC. e Genome editing of the RIG-I gene in CHSE-EC. CHSE-EC cells were transduced similar to EGFP targeting (Puro-) and selected with puromycin (Puro+) and efficiency estimated by ICE analysis of Sanger sequencing chromatogram from the PCR amplified target region. f Detail of indels frequency in RIG-I edited Puro- group (from panel e), estimated by ICE analysis. Purple dot denotes the unedited sequence (0 bp)