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Flp-In T-REx 293 cells stably integrated with an inducible 3xFLAG-HERC2 construct (WT), its catalytically inactive mutant (C4762S), or empty vector as a control (-) were treated with doxycycline to induce the expression of each HERC2 construct (seeFigure S3E). Cytosolic extracts (input) were separated on a 5%–45% sucrose gradient and analyzed by immunoblotting for CCT4 and CCT7. Unassembled CCT subunits migrate in fractions 2–4, whereas assembled CCT migrates in fractions 7–9.

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