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Article|19 Jan 2022|OPEN
The callus formation capacity of strawberry leaf explants is modulated by DNA methylatio
Decai Liu1 , Qin Mu1 , Xianyang Li1 , Sheng Xu1 , Yi Li2 , and Tingting Gu,1 ,
1State Key Laboratory of Plant Genetics and Germplasm Enhancement and College of Horticulture, Nanjing Agricultural University, Nanjing, 210095 China
2Department of Plant Science and Landscape Architecture, University of Connecticut, Storrs, CT 06269, USA
*Corresponding author. E-mail:,

Horticulture Research 9,
Article number: uhab073 (2022)
Views: 24

Received: 08 Aug 2021
Accepted: 04 Nov 2021
Published online: 19 Jan 2022


Shoot regeneration from leaf tissue requires the de-differentiation of cells from a highly differentiated state into an actively dividing state, but it remains unclear how this physiological transition occurs and is regulated, especially at the epigenetic level. Here, we characterized the DNA methylome represented by 5-methylcytosine (5mC) in leaf and callus tissue derived from leaf explants of woodland strawberry, Fragaria vesca. We detected an overall increase in DNA methylation and distinct 5mC enrichment patterns in the CG, CHG, and CHH sequence contexts in genes and transposable elements. Our analyses revealed an intricate relationship between DNA methylation and gene expression level in leaves or leaf-derived callus. However, when considering the genes involved in callus formation and shoot regeneration, e.g. FvePLT3/7FveWIND3FveWIND4FveLOG4 and FveIAA14, their dynamic transcription levels were associated with differentially methylated regions located in the promoters or gene bodies, indicating a regulatory role of DNA methylation in the transcriptional regulation of pluripotency acquisition in strawberry. Furthermore, application of the DNA methyltransferase inhibitor 5′-azacytidine (5′-Aza) hampered both callus formation and shoot regeneration from the leaf explants. We further showed that 5′-Aza downregulated the expression of genes involved in cell wall integrity, such as expansinpectin lyase, and pectin methylesterase genes, suggesting an essential role of cell wall metabolism during callus formation. This study reveals the contribution of DNA methylation to callus formation capacity and will provide a basis for developing a strategy to improve shoot regeneration for basic and applied research applications.