Gaining a mechanistic understanding of the molecular pathways underpinning cellular and organismal physiology invariably relies on the perturbation of an experimental system to infer causality. This can be achieved either by genetic manipulation or by pharmacological treatment. Generally, the former approach is applicable to a wider range of targets, is more precise, and can address more nuanced functional aspects. Despite such apparent advantages, genetic manipulation (i.e., knock‐down, knock‐out, mutation, and tagging) in mammalian systems can be challenging due to problems with delivery, low rates of homologous recombination, and epigenetic silencing. The advent of CRISPR‐Cas9 in combination with the development of robust differentiation protocols that can efficiently generate a variety of different cell typesin vitrohas accelerated our ability to probe gene function in a more physiological setting. Often, the main obstacle in this path of enquiry is to achieve the desired genetic modification. In this short review, we will focus on gene perturbation in mammalian cells and how editing and differentiation of pluripotent stem cells can complement more traditional approaches. Additionally, we introduce novel targeted protein degradation approaches as an alternative to DNA/RNA‐based manipulation. Our aim is to present a broad overview of recent approaches andin vitrosystems to study mammalian cell biology. Due to space limitations, we limit ourselves to providing the inexperienced reader with a conceptual framework on how to use these tools, and for more in‐depth information, we will provide specific references throughout.