Synchronization based on precise cell cycle progression identification to enhance tetraploid induction in '84 K' poplar.

IF 4.5 2区 生物学 Q1 PLANT SCIENCES
Aoyu Ling, Yufei Xia, Yijia Jin, Shenxiu Jiang, Jianghai Shu, Kang Du, Pingdong Zhang, Xiangyang Kang
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Abstract

Key message: This research develops a novel method for improving tetraploid induction efficiency in vitro by precisely identifying key cell cycle phases and optimizing synchronization treatments in '84 K' poplar. Cell cycle asynchrony is a critical bottleneck in tetraploid in vitro induction efficiency. 5-Aminouracil (5-AU), a known cell cycle inhibitor, provides a strategy to enhance tetraploid induction efficiency. However, as 5-AU primarily targets S-phase cells and demonstrates cytotoxicity, improving tetraploid induction efficiency requires first accurately identifying the peak window of S-phase cell accumulation in the cultured tissue. Additionally, it is crucial to determine the appropriate concentration range of 5-AU to minimize toxicity to the cultured tissue. In this study, we developed a cell cycle synchronization strategy and evaluated its effect on tetraploid induction efficiency. Flow cytometry and EdU fluorescence labeling were used to track the cell cycle progression of '84 K' poplar (Populus alba × P. glandulosa) leaves during in vitro culture. Our results showed that S-phase activity peaked between days 2 and 3 of differentiation, while the G2/M phase began on day 4. This defined the optimal synchronization window as days 2 to 3. Comet assays were conducted to evaluate DNA damage induced by 5-AU. The results showed that DNA fragmentation increased significantly at concentrations exceeding 1.7 mM. In contrast, at concentrations below 1.3 mM, comet assays indicated relatively low DNA damage, while flow cytometry analysis revealed suboptimal synchronization efficiency. Consequently, 1.5 mM was identified as the optimal concentration for synchronization induction. By applying this synchronization strategy, the tetraploid induction rate of '84 K' poplar was increased to an average of 40.90%. Our results demonstrate that precise synchronization of the cell cycle, with minimal cytotoxicity, can significantly improve tetraploid induction efficiency.

基于细胞周期进程精确鉴定的同步提高84k杨树四倍体诱导。
本研究提出了一种新的方法,通过精确识别关键细胞周期期和优化同步处理,提高‘84 K’杨树四倍体体外诱导效率。细胞周期不同步是影响四倍体体外诱导效率的关键瓶颈。5-氨基尿嘧啶(5-AU)是一种已知的细胞周期抑制剂,提供了提高四倍体诱导效率的策略。然而,由于5-AU主要靶向s期细胞并具有细胞毒性,因此提高四倍体诱导效率首先需要准确识别培养组织中s期细胞积累的峰值窗口。此外,确定合适的5-AU浓度范围以尽量减少对培养组织的毒性也是至关重要的。在本研究中,我们开发了一种细胞周期同步策略,并评估了其对四倍体诱导效率的影响。采用流式细胞术和EdU荧光标记技术对84 K杨树(Populus alba × P)细胞周期进行了跟踪研究。离体培养期间的腺体)叶。结果表明,s期活性在分化第2 ~ 3天达到高峰,而G2/M期开始于分化第4天。这将最佳同步窗口定义为第2天到第3天。采用彗星法评价5-AU诱导的DNA损伤。结果表明,当浓度超过1.7 mM时,DNA片段明显增加。相反,当浓度低于1.3 mM时,彗星分析显示DNA损伤相对较低,而流式细胞术分析显示同步效率不佳。因此,1.5 mM为同步诱导的最佳浓度。采用该同步策略,84 K杨树四倍体诱导率平均提高到40.90%。我们的研究结果表明,细胞周期的精确同步,以最小的细胞毒性,可以显著提高四倍体诱导效率。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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来源期刊
Plant Cell Reports
Plant Cell Reports 生物-植物科学
CiteScore
10.80
自引率
1.60%
发文量
135
审稿时长
3.2 months
期刊介绍: Plant Cell Reports publishes original, peer-reviewed articles on new advances in all aspects of plant cell science, plant genetics and molecular biology. Papers selected for publication contribute significant new advances to clearly identified technological problems and/or biological questions. The articles will prove relevant beyond the narrow topic of interest to a readership with broad scientific background. The coverage includes such topics as: - genomics and genetics - metabolism - cell biology - abiotic and biotic stress - phytopathology - gene transfer and expression - molecular pharming - systems biology - nanobiotechnology - genome editing - phenomics and synthetic biology The journal also publishes opinion papers, review and focus articles on the latest developments and new advances in research and technology in plant molecular biology and biotechnology.
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