Seeds are an important life cycle stage because they guarantee plant survival in unfavorable environmental conditions and the transfer of genetic information from parents to offspring. However, similar to every organ, seeds undergo aging processes that limit their viability and ultimately cause the loss of their basic property, i.e., the ability to germinate. Seed aging is a vital economic and scientific issue that is related to seed resistance to an array of factors, both internal (genetic, structural, and physiological) and external (mainly storage conditions: temperature and humidity). Reactive oxygen species (ROS) are believed to initiate seed aging via the degradation of cell membrane phospholipids and the structural and functional deterioration of proteins and genetic material. Researchers investigating seed aging claim that the effective protection of genetic resources requires an understanding of the reasons for senescence of seeds with variable sensitivity to drying and long-term storage. Genomic integrity considerably affects seed viability and vigor. The deterioration of nucleic acids inhibits transcription and translation and exacerbates reductions in the activity of antioxidant system enzymes. All of these factors significantly limit seed viability.

Background and Aims Epigenetic regulation plays an important role in the management of plant growth, development and response to stress factors, and several reports have indicated that DNA methylation plays a critical role in seed development and viability. This study examines changes in 5-methylcytosine (m⁵C) levels in the DNA of seeds during ageing, a process that has important implications for plant conservation and agriculture. Methods Changes in the global level of m⁵C were measured in mature seeds of oak, Quercus robur. The extent of DNA methylation was measured using a protocol based on two-dimensional thin-layer chromatography. Viability of seeds was determined by germination and seedling emergence tests. Key Results An ageing-related decrease in total m⁵C during storage of recalcitrant seeds was highly and significantly correlated with a decrease in seed viability, as reflected by a reduction in germination (r = 0·8880) and seedling emergence (r = 0·8269). Conclusions The decrease in viability during ageing of Q. robur seeds is highly correlated with a global decline in the amount of m⁵C in genomic DNA, and it is possible that this may represent a typical response to ageing and senescence in recalcitrant seeds. Potential mechanisms that drive changes in genomic DNA methylation during ageing are discussed, together with their implications for seed viability.

Seeds are the most commonly used source of storage material to preserve the genetic diversity of plants. However, prior to the deposition of seeds in gene banks, several questions need to be addressed. Here, we illustrate the scheme that can be used to ensure that the most optimal conditions are identified to enable the long-term storage of seeds. The main questions that need to be answered pertain to the production of viable seeds by plants, the availability of proper protocols for dormancy alleviation and germination, seed tolerance to desiccation and cold storage at −20 °C. Finally, it is very important to fully understand the capability or lack thereof for seeds or their explants to tolerate cryogenic conditions. The proper storage regimes for orthodox, intermediate and recalcitrant seeds are discussed.

Poor storability of recalcitrant seeds is due to their inability to tolerate low moisture content. Understanding the processes underlying their recalcitrance is a prerequisite to developing a maintenance strategy and prolonging their lifespan. Multiple studies have investigated the differences between orthodox (desiccation-tolerant) and recalcitrant (desiccation-sensitive) seeds. Information on epigenetic regulation, however, is lacking and thus limits our understanding of the processes defining the physiology of seeds. In the present comparative study, changes in the global levels of 5-methylcytosine (m⁵C) in orthodox and recalcitrant seeds of Acer platanoides L. and Acer pseudoplatanus L. were characterized during progressive stages of severe drying. Concomitant with their differential sensitivity to desiccation stress, we demonstrate variation in the response of embryonic axes and cotyledons to water deficit at the level of DNA methylation. Results indicate that desiccation-induced changes in m⁵C are both tissue- and seed category-specific and are highly correlated with recalcitrant seed viability. Moreover, we demonstrate that m⁵C global changes in response to desiccation are not retained in DNA isolated from seedlings, except in seedlings that are derived from strongly desiccated orthodox seeds (moisture content of 3.5%). Finally, the potential utilization of m⁵C status as a universal seed viability marker is discussed.

Modifications of DNA nucleobases are present in all forms of life. The purpose of these modifications in eukaryotic cells, however, is not always clear. Although the role of 5-methylcytosine (m⁵C) in epigenetic regulation and the maintenance of stability in plant genomes is becoming better understood, knowledge pertaining to the origin and function of oxidized nucleobases is still scarce. The formation of 5-hydroxymetylcytosine (hm⁵C) in plant genomes is especially debatable. DNA modifications, functioning as regulatory factors or serving as DNA injury markers, may have an effect on DNA structure and the interaction of genomic DNA with proteins. Thus, these modifications can influence plant development and adaptation to environmental stress. Here, for the first time, the changes in DNA global levels of m⁵C, hm⁵C, and 8-oxo-7,8-dihydroguanine (8-oxoG) measured by ELISA have been documented in recalcitrant embryonic axes subjected to desiccation and accelerated aging. We demonstrated that tissue desiccation induces a similar trend in changes in the global level of hm⁵C and 8-oxoG, which may suggest that they both originate from the activity of reactive oxygen species (ROS). Our study supports the premise that m⁵C can serve as a marker of plant tissue viability whereas oxidized nucleobases, although indicating a cellular redox state, cannot.