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Despite broad scientific interest in harnessing the power of Earth’s microbiomes, knowledge gaps hinder their efficient use for addressing urgent societal and environmental challenges. We argue that structuring research and technology developments around a design– build–test–learn (DBTL) cycle will advance microbiome engineering and spur new discoveries of the basic scientific principles governing microbiome function. In this Review, we present key elements of an iterative DBTL cycle for microbiome engineering, focusing on generalizable approaches, including top- down and bottom- up design processes, synthetic and self- assembled construction methods, and emerging tools to analyse microbiome function. These approaches can be used to harness microbiomes for broad applications related to medicine, agriculture, energy and the environment. We also discuss key challenges and opportunities of each approach and synthesize them into best practice guidelines for engineering microbiomes. We anticipate that adoption of a DBTL framework will rapidly advance microbiome- based biotechnologies aimed at improving human and animal health, agriculture and enabling the bioeconomy. 

Oxygen levels in vivo are autonomously regulated by a supply–demand balance, which can be altered in disease states. However, the oxygen levels of in vitro cell culture systems, particularly microscale cell culture, are typically dominated by either supply or demand. Further, the oxygen microenvironment in these systems is rarely monitored or reported. Here, a method to establish and dynamically monitor autonomously regulated oxygen microenvironments (AROM) using an oil overlay in an open microscale cell culture system is presented. Using this method, the oxygen microenvironment is dynamically regulated via the supply–demand balance of the system. Numerical simulation and experimental validation of oxygen transport within multi‐liquid‐phase, microscale culture systems involving a variety of cell types, including mammalian, fungal, and bacterial cells are presented. Finally, AROM is applied to establish a coculture between cells with disparate oxygen demands—primary intestinal epithelial cells (oxygen consuming) andBacteroides uniformis(an anaerobic species prevalent in the human gut).