Digital sight ds fi1 camera system

The Surveyor mutagenesis assay was used to validate activity of some vectors (Figure S3). OSE cells were transduced with Trp53-targeting LentiCRISPRs because OSN2 cells lack Trp53 alleles. OSN2 cells were transduced with redesigned LentiCRISPRs intended to replace non-functional constructs. Briefly, LentiCRISPR target sites were PCR-amplified in transduced (edited) and untransduced (control) OSE cells (Supplemental Table 2). Amplicons from control and edited cells were mixed in Transformation was monitored for one week, and colonies were collected via the manufacturer's instructions. In short, Cell Biolabs Inc matrix solubilization solution was added to each well of cells (Cell Biolabs; Cat#: CBA-135). Colonies were resuspended in OSE media and plated at very low density on 15cm plates. Individual, distinct colonies growing on 15cm plates were picked and cultured independently. Sterile filter paper was soaked in trypsin and was used to pick individual colonies from plates.
Picked colonies were added to 24-well plates and were passaged using OSE culturing methodology. Brightfield images of culture wells were taken using a Nikon SMZ1500 microscope and Nikon Digital Sight DS-Fi1 camera system at 2x magnification. 10x and 20x images were taken using a Nikon TMS-F microscope and Moticam 2300 3.0MP Live Resolution camera system.