For the experiment in Fig 1D , mouse ES cells expressing DONSON‐GFP and mCherry‐PSF1 from the endogenous loci were grown on “μ‐Slide 4‐well” (Ibidi, 80426) with “no phenol red DMEM medium” (ThermoFisher Scientific, 21063029) supplemented as described above. For p97 inhibition, cells were treated with 5 μM CB‐5083 for 3 h before imaging.
Confocal images of live cells were acquired with a Zeiss Cell Observer SD microscope with a Yokogawa CSU‐X1 spinning disk, using a HAMAMATSU C13440 camera with a PECON incubator, a 60× 1.4‐NA Plan‐Apochromat oil immersion objective, and excitation and emission filter sets for GFP and mCherry. Images of live mouse ES cells were acquired using “ZEN blue” software (Zeiss) and processed with ImageJ software (National Institutes of Health) as previously described (Sonneville et al, 2017 (link)).
Confocal images of live cells were acquired with a Zeiss Cell Observer SD microscope with a Yokogawa CSU‐X1 spinning disk, using a HAMAMATSU C13440 camera with a PECON incubator, a 60× 1.4‐NA Plan‐Apochromat oil immersion objective, and excitation and emission filter sets for GFP and mCherry. Images of live mouse ES cells were acquired using “ZEN blue” software (Zeiss) and processed with ImageJ software (National Institutes of Health) as previously described (Sonneville et al, 2017 (link)).
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