A newly identified gene boosts wheat regeneration and transformation efficiency

By directly suppressing cytokinin degradation genes (TaCKXs) and promoting key developmental genes (TaWOX5 and TaGIF1), TaDOF4.7-B dramatically increases shoot regeneration and callus proliferation.

Wheat is a globally vital food crop, yet its limited regeneration ability has long hampered genetic transformation and molecular breeding efforts. Plant regeneration—the ability to develop new organs or tissues—is a cornerstone of modern crop biotechnology, enabling genetic transformation and variety improvement. Among plant hormones, cytokinin plays a central role in stimulating callus formation and shoot development. Its homeostasis is tightly regulated by synthesis enzymes (IPTs) and degradation enzymes (CKXs). While manipulating cytokinin levels has shown promise in improving regeneration, the transcriptional mechanisms that coordinate this balance in wheat remain poorly understood. Based on these challenges, identifying key transcription factors that control cytokinin homeostasis is essential for advancing wheat regeneration and transformation efficiency.

A study (DOI: 10.48130/seedbio-0025-0018 ) published in Seed Biology on 03 November 2025 by Haixia Yu’s & Chen Wang’s team, Shandong Agricultural University, provides crucial insights into the molecular mechanisms underlying plant regeneration and paves the way for more efficient genome editing and variety improvement in wheat.

To investigate the role of the TaDOF4.7-B transcription factor in wheat regeneration, several experimental techniques were employed. First, chromatin accessibility analysis identified that the promoters of TaDOF4.7-B and TaDOF4.7-D had higher chromatin accessibility compared to TaDOF4.7-A during the early stages of callus induction. RT-qPCR confirmed that TaDOF4.7-B was more highly expressed than the other DOF family members during wheat regeneration. To explore its function, TaDOF4.7-B was fused with green fluorescent protein (GFP) and expressed in Nicotiana benthamiana, revealing its nuclear localization. In wheat, RNA in-situ hybridization showed TaDOF4.7-B was primarily expressed in regions of callus where shoot primordia would emerge. Further, overexpression of TaDOF4.7-B in Fielder wheat led to earlier shoot primordia formation, better callus growth, and higher regeneration frequencies. Specifically, regeneration frequency increased from 22.65% in controls to 60.57% in transgenic lines, while regenerating shoot frequency and callus proliferation also significantly improved. Next, transcriptome analysis was performed on calli collected at different regeneration stages. Overexpression of TaDOF4.7-B resulted in the upregulation of genes related to cell division, shoot formation, and cytokinin signaling, including TaWOX5 and TaGIF1. ChIP-qPCR demonstrated that TaDOF4.7-B directly binds to the promoters of these genes, suggesting it regulates their expression during shoot regeneration. The overexpression lines also showed altered cytokinin levels, as the expression of TaCKX genes—responsible for cytokinin degradation—was downregulated. This regulation of cytokinin homeostasis by TaDOF4.7-B significantly enhanced shoot regeneration, even in the absence of exogenous cytokinin. These results highlight the critical role of TaDOF4.7-B in wheat regeneration and provide valuable insights for improving wheat transformation efficiency.

The discovery of TaDOF4.7-B offers a new genetic tool for improving transformation efficiency in wheat, a major bottleneck in cereal biotechnology. By modulating cytokinin metabolism rather than relying on hormone supplementation, this approach enables faster and more reliable regeneration across genotypes. Integrating TaDOF4.7-B into transformation pipelines could accelerate the production of gene-edited or transgenic wheat varieties with improved yield, quality, and stress tolerance. Beyond wheat, this regulatory mechanism provides a model for enhancing regeneration in other recalcitrant crops, offering broad applications in plant breeding and agricultural biotechnology.

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References

DOI

10.48130/seedbio-0025-0018

Original Source URL

https://doi.org/10.48130/seedbio-0025-0018

Funding information

This research was funded by the Biological Breeding-National Science and Technology Major Project (2024ZD04077).

About Seed Biology

Seed Biology (e-ISSN 2834-5495) is published by Maximum Academic Press in partnership with Yazhou Bay Seed Laboratory. Seed Biology is an open access, online-only journal focusing on research related to all aspects of the biology of seeds, including but not limited to: evolution of seeds; developmental processes including sporogenesis and gametogenesis, pollination and fertilization; apomixis and artificial seed technologies; regulation and manipulation of seed yield; nutrition and health-related quality of the endosperm, cotyledons, and the seed coat; seed dormancy and germination; seed interactions with the biotic and abiotic environment; and roles of seeds in fruit development. Seed biology publishes a wide range of research approaches, such as omics, genetics, biotechnology, genome editing, cellular and molecular biology, physiology, and environmental biology. Seed Biology publishes high-quality original research, reviews, perspectives, and opinions in open access mode, promoting fast submission, review, and dissemination freely to the global research community.