Somatic embryogenesis is a phenomenon carried out in an environment that generates abiotic stress. Thus, regenerants may differ from the source of explants at the morphological, genetic, and epigenetic levels. The DNA changes may be the outcome of induction media ingredients (i.e., copper and silver ions) and their concentrations and time of in vitro cultures. This study optimised the level of copper and silver ion concentration in culture media parallel with the induction medium longevity step towards obtaining barley regenerants via somatic embryogenesis with a minimum or maximum level of tissue culture-induced differences between the donor plant and its regenerants. The optimisation process is based on tissue culture-induced variation evaluated via the metAFLP approach for regenerants derived under varying in vitro tissue culture conditions and exploited by the Taguchi method. In the optimisation and verification experiments, various copper and silver ion concentrations and the different number of days differentiated the tested trials concerning the tissue culture-induced variation level, DNA demethylation, and de novo methylation, including symmetric (CG, CHG) and asymmetric (CHH) DNA sequence contexts. Verification of optimised conditions towards obtaining regenerants with minimum and maximum variability compared to donor plants proved useful. The main changes that discriminate optimised conditions belonged to DNA demethylation events with particular stress on CHG context. The combination of tissue culture-induced variation evaluated for eight experimental trials and implementation of the Taguchi method allowed the optimisation of the in vitro tissue culture conditions towards the minimum and maximum differences between a source of tissue explants (donor plant) and its regenerants from somatic embryos. The tissue culture-induced variation characteristic is mostly affected by demethylation with preferences towards CHG sequence context.

中文翻译：

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大麦体细胞胚发生-试图改变组织培养中诱导的变异

体细胞胚发生是在产生非生物胁迫的环境中进行的现象。因此，在形态，遗传和表观遗传水平上，再生子可能与外植体的来源不同。DNA的变化可能是诱导培养基成分（即铜和银离子）及其在体外培养物中的浓度和时间的结果。这项研究优化了培养基中铜和银离子的浓度水平，并通过诱导诱导体寿命的步骤，通过体细胞胚发生获得了大麦再生体，同时在供体植物及其再生体之间组织培养物诱导的差异达到了最小或最大水平。优化过程基于通过组织培养诱导的变异，该变异通过metAFLP方法评估了在不同体外组织培养条件下衍生并由Taguchi方法开发的再生体。在优化和验证实验中，各种铜和银离子浓度和不同天数使有关组织培养物诱导的变异水平，DNA去甲基化和从头甲基化（包括对称（CG，CHG）和不对称（ CHH）DNA序列背景。与供体植物相比，验证优化条件以获得具有最小和最大变异性的再生体被证明是有用的。区分优化条件的主要变化是DNA脱甲基事件，尤其是CHG环境。通过对组织培养物诱导的变异进行了八项实验试验评估和Taguchi方法的实施，可以优化体外组织培养条件，以实现组织外植体来源（供体植物）与其体细胞再生体之间的最小和最大差异。胚胎。组织培养物诱导的变异特征主要受到去甲基化的影响，偏爱CHG序列。