Tandem affinity purification of functional TAP-tagged proteins from human cells |
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| Authors: | Gregan Juraj Riedel Christian G Petronczki Mark Cipak Lubos Rumpf Cornelia Poser Ina Buchholz Frank Mechtler Karl Nasmyth Kim |
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| Affiliation: | Department of Chromosome Biology, Max F. Perutz Laboratories, University of Vienna, Dr Bohr-Gasse 1, 1030 Vienna, Austria. juraj.gregan@univie.ac.at |
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| Abstract: | Tandem affinity purification (TAP) is a generic two-step affinity purification protocol for isolation of TAP-tagged proteins together with associated proteins. We used bacterial artificial chromosome to heterologously express TAP-tagged murine Sgo1 protein in human HeLa cells. This allowed us to test the functionality of the Sgo1-TAP protein by RNA interference-mediated depletion of the endogenous human Sgo1. Here, we present an optimized protocol for purification of TAP-tagged Sgo1 protein as well as KIAA1387 from HeLa cells with detailed instructions. The purification protocol can be completed in 1 day and it should be applicable to other proteins. |
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