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Wild-type or single-lysine HA-tagged ubiquitin was expressed in HEK-293T cells. Ubiquitinated proteins were immunoprecipitated with anti-HA antibodies and blotted with anti–K63-specific or anti–K48-specific antibodies as well as antibodies against pan-ubiquitin. Note that both antibodies and single-lysine ubiquitin constructs have the expected specificity without any cross-reactivity detected.
