Studies of the gut microbiota have revealed associations between specific bacterial species or community compositions with health and disease, yet the causal mechanisms underlying microbiota gene–host interactions remain poorly understood. This is partly due to limited genetic manipulation (GM) tools for gut bacteria. Here, we review current advances and challenges in the development of GM approaches, including CRISPR-Cas and transposase-based systems in either model or non-model gut bacteria. By overcoming barriers to ‘taming’ the gut microbiome, GM tools allow molecular understanding of host–microbiome associations and accelerate microbiome engineering for clinical treatment of cancer and metabolic disorders. Finally, we provide perspectives on the future development of GM for gut microbiome species, where more effort should be placed on assembling a generalized GM pipeline to accelerate the application of groundbreaking GM tools in non-model gut bacteria towards both basic understanding and clinical translation.