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Assembly and use of a plant gnotobiotic growth system for plant–microbe interaction studies in cereals

Tang, Clara1; Pal, Gaurav1; Ring, Nathan1; Pietrosemoli, Daniel2; Grunden, Amy1; Kleiner, Manuel1

Published Apr 07, 2026 on Dryad. https://doi.org/10.5061/dryad.f7m0cfzbc

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Abstract

The development of microbial-based agricultural amendments that work consistently in the field requires an understanding of the molecular mechanisms of plant–microbe interactions. Studying these underlying mechanisms of interaction demands the ability to grow plants under environmentally controlled and gnotobiotic conditions (i.e. all microorganisms interacting with the plant are known, whether that is germ-free, defined microbial communities, or natural communities). The currently available plant gnotobiotic systems are not suitable to study large plants of agricultural relevance such as cereals. Moreover, most of these systems lack the ability to manage irrigation.  Here we introduce GNOVA, a new gnotobiotic system designed to accommodate cereal plants with the ability to manage irrigation. This  new system is an accessible platform composed of a 3D printed base and commercially available materials. This protocol provides a step-by-step guide to assembling the system and experiment set up. Furthermore, we present a performance comparison of GNOVA to a gnotobiotic bag system. GNOVA extended plant growth from two weeks in the bag system up to 17 weeks for wheat and 4 weeks for maize. The germination rate of both crops also increased within GNOVA from 66% to 100% for wheat and from 75% to 100% for maize. Wheat grown within GNOVA developed tillers, which were absent in plants of the same age within the bag system. The fresh weight of maize grown in the GNOVA was 594% higher than in the bag system. Additionally, the shoot height and root length of maize were 89% and 57% greater within the GNOVA system than in the bags, respectively. The GNOVA system extends the toolbox available to scientists for the exploration of plant–microbiota interactions beyond the seedling stage in cereals by providing increased growth space and irrigation management.