Impact of Rejuvenation Measures on Growth and Tissue Culture Regeneration of Fraxinus mandshurica

doi:10.7525/j.issn.1673-5102.2026.01.008

Abstract

Abstract:

As an essential method for clonal propagation of the valuable timber species Fraxinus mandshurica， tissue culture is hindered by physiological aging of explants due to age-related effects， which significantly reduces tissue culture efficiency and severely restricts the rapid clonal propagation and widespread application of superior F. mandshurica varieties. Currently， there are no reports on overcoming the age effect through rejuvenation to improve tissue culture efficiency in forest trees. This study investigated the effects of different rejuvenation treatments on growth performance and tissue culture regeneration of 4-year-old F. mandshurica plants. The results showed that after trunk cutting + shading treatment， the height and diameter of new shoots decreased by 37.50% and 42.90%， respectively， while the number of new shoots and stem segment explants increased by 2.6-fold and 2.8-fold， which significantly enhanced tissue culture regeneration capacity. The initiation rate of stem segment explants， new shoot differentiation rate， and propagation coefficient of the shoots under treatment all increased compared to the untreated control（CK）. The highest initiation rate reached 82.42%， which was 3.26 times as high as that of the CK（25.30%）. The differentiation rate increased from 9.90% to 61.11%. The propagation coefficient increased from 0 to 6.00. Further investigation into the mechanism by which rejuvenation measures overcome the age effect was conducted through the analysis of physiological and biochemical indicators and related gene expression changes. It was found that after rejuvenation treatment， sucrose and soluble sugar content significantly decreased by 23.17% and 46.24%， while the content of auxin（IAA） and cytokinin（CTK） significantly increased by 70.01% and 57.22%， respectively. Based on the trend of miR156A/C gene expression downregulating with plant maturation， transcriptional analysis of the miR156-SPL-AP2/ERFs-ASA1 gene pathway further revealed that after trunk cutting + shading treatment， the expression levels of rejuvenation and bud regeneration-related genes FmmiR156A， FmmiR156C， and FmASA1 increased by 10.01， 13.53， and 12.58 times compared to CK， while the expression levels of FmSPL2 and FmSPL9 decreased by 93.05% and 88.30% compared to CK. These findings indicated that the above indicators may be key factors promoting rejuvenation in F. mandshurica， thereby improving the tissue culture regeneration efficiency of explants. This study provided practical methods for addressing the age effect and improving tissue culture propagation capacity in F. mandshurica and other forest trees species， offering technical support for the large-scale propagation and promotion of superior varieties and new cultivars of F. mandshurica.

Key words: Fraxinus mandshurica, tissue culture, age effect, rejuvenation treatment, physiological and biochemical indicators, gene expression

CLC Number:

Q945.4

Chengming XIONG, Qiaoxin WANG, Liming HE, Jianfei LIU, Guangzhou PENG, Anhong HU, Xinyu LI, Yaguang ZHAN. Impact of Rejuvenation Measures on Growth and Tissue Culture Regeneration of Fraxinus mandshurica[J]. Bulletin of Botanical Research, 2026, 46(1): 91-100.

Figures/Tables 6

Table 1

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不同复幼处理 Different rejuvenation treatments	外植体启动率 Explant initiation rate/%	芽分化率 Differentiation rate/%	分化芽数量 Number of differentiated buds	增殖系数 Propagation coefficient	隶属函数值 D	排名 Ranking
截干+遮阴 Trunk cutting+shading	82.42	61.11	5.67	6.00	0.99	1
循环3次 Three cycles of rejuvenation	82.59	63.33	6.33	6.63	0.76	2
截干 Trunk cutting	74.03	44.44	4.33	4.23	0.76	3
埋干 Stem layering for shoot regeneration	85.74	25.56	1.33	3.77	0.63	4
循环2次 Two cycles of rejuvenation	70.37	55.56	4.67	5.87	0.61	5
循环1次 One cycle of rejuvenation	56.30	34.44	2.67	4.43	0.41	6
未处理 CK	25.30	9.90	1.33	0	0.09	7