The organization of the genome inside the nucleus facilitates many nuclear processes. Because the nuclear genome is highly dynamic and often regulated by essential proteins, rapid depletion strategies are necessary to perform loss-of-function analyses. Fortunately, in recent years, various methods have been developed to manipulate the cellular levels of a protein directly and acutely. Here, we describe different methods that have been developed to rapidly deplete proteins from cells, with a focus on auxin inducible degron and dTAG methods, as these are most commonly used in 3D genome organization studies. We outline best practices for designing a knockin strategy, as well as generation and validation of knockin cell lines. Acute depletion strategies have been transformative for the study of the 3D genome and will be important tools for delineating the processes and factors that determine organization of the genome inside the nucleus.
Keywords: Acute protein degradation; Chromatin dynamics; Genome editing; Genome organization; Protein knockdown.
© 2022. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.