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bioRxiv · 10.64898/2026.01.20.700461

A conserved and predictable pluripotency window in callus unlocks efficient transformation in grasses and beyond

Wang, Y.·Chu, M.·Wang, Z.·Shao, J.·Zhang, H.·Nan, Z.·Li, C.·Lei, L.
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Abstract

A major bottleneck in plant biotechnology is the inefficient and genotype-dependent regeneration of callus, which severely limits genetic transformation and functional studies across many species. This barrier is acutely exemplified in the study of beneficial plant-microbe interactions, such as the Epichloe-grass symbiosis--a system conferring remarkable stress tolerance to its host but hindered by a lack of efficient genetic tools. To address this, we established a chromosome-scale genome for an Epichloe native host grass Achnatherum inebrians. We discovered that the expression dynamics of evolutionarily conserved cell pluripotency regulators (CPRs) including ARF5/7/19, BBM, WUS/WOX5 and CUC1/2 serve as a precise molecular predictor for callus regenerative capacity, revealing that pluripotency is dynamic and peaks within a narrow, definable time window. Harnessing this predictable window enabled the development of a highly efficient transformation system for A. inebrians (49.4% efficiency). Crucially, this CPR-based strategy proved generalizable: applied to wheat and the legume sainfoin, it pinpointed species-specific optimal regeneration windows, boosting shoot regeneration rates to 65.7% and 87.5%, respectively. Collectively, our work provides an integrated research system and a rational design principle that removes a key barrier to uncovering molecular mechanisms in plant systems, particularly the Epichloe-enhanced stress tolerance symbiosis.

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Wang, Y., Chu, M., Wang, Z., Shao, J., Zhang, H., Nan, Z., Li, C., Lei, L.. 2026-01-23. A conserved and predictable pluripotency window in callus unlocks efficient transformation in grasses and beyond. https://doi.org/10.64898/2026.01.20.700461

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