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1/4 -Example of in-Site and SCREEN combination assay in unedited human fibroblast control line cell. Bracketing in-Site probes on chromosome 22 (circled) to identify insert target location and dGH SCREEN paints on chromosomes 1, 2, and 3 (pink). No insert would be expected in this control line
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2/4 - Example of a single cell from the same in-Site assay in an edited iPSC viewed in multiple fluorescence channels. Left image shows all fluorescence channels overlaid, middle shows yellow/insert probe channel, and right shows green/on-target bracketing probe channel. Multiple inserts are visible outside of the on-target bracketing probes, indicating high levels of off-target integrations in this cell.
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3/4 - Example of the same in-Site assay in an edited iPSC line cell with no off target insertions. Target site bracketing probes in green and insert probe in yellow zoomed in and expanded on the right. Results from this sample indicated that 6% of cells had no off-target insertions.
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4/4 - Image from a dGH in-Site assay in polyploid, dual-edited, HEK-293FT cells demonstrating both normal and rearranged edit site structures. At the normal edit sites (A), both sets of in-Site probes are located on the same side of the chromosome. An inversion at the edit site is also visible (B), with one portion of the in-Site probe switching orientations and showing up on the opposite chromatid from the other portions of the in-Site probe. A partial deletion at one edit site is also observed (C), with the red portion of the in-Site probes missing.