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Rejuvenation is an efficient approach used in the cuttings of trees and horticultural crops, to improve their rooting ability, especially in difficult-to-root trees. WOX gene family members are involved in cell-fate transformation through balancing the maintenance and proliferation of the stem cells. However, there are no reports about the WOX gene family in Walnut (Juglans regia L.) and its relationship between rejuvenation and adventitious roots formation (ARF). Here, a genome-wide identification of JrWOX genes and their physical and chemical properties, phylogeny, and expression profiles in different organs and during rejuvenation-induced ARF is reported. The phenotype and histology characteristics of mature and rejuvenated cuttings (Mc and Rc) are also observed. In this study, 12 genes were identified and clustered into three groups based on phylogenetics, special domains, and conserved motifs. The gene structures and conserved motifs were relatively conserved, while the 12 sequences of the JrWOXs domain were diversified. Gene expression in root, stem, leaf, female flower, immature fruit, and zygotic embryo revealed that the expression levels of JrWOX4a, JrWOX4b, JrWOX5, JrWOX11, and JrWOX13 in the root were significantly higher than those of other JrWOXs, while only the expression of JrWOX11 was exclusive to the root organ. Additionally, rejuvenation treatment significantly induced almost all JrWOX genes, except JrWOX4a, JrWOX4b, and JrWOX13 (Rc 0 vs. Mc 0). During the ARF process, the transcripts of JrWOX11 and JrWOX5 were consecutively increased on a significance level; in contrast, the transcription levels of the other JrWOXs decreased or changed insignificantly. The phenotype and histology observation indicate that rejuvenation treatment made the base of the stem expand and reduced the thickness and density of sclerenchyma between the cortex and phloem. This might provide the conditions for the formation of new meristem niches. The results provided insight into the JrWOX genes’ general characteristics and their roles in rejuvenation-induced ARF.

Genome-Wide Identification of WOX Gene Family and Expression Analysis during Rejuvenational Rhizogenesis in Walnut (Juglans regia L.)