中国科学院分子植物科学卓越创新中心/中国科学院植物生理生态研究所王佳伟团队报道了一种对难以消化和冷冻保存的植物样本进行单细胞RNA测序的方法。这一研究成果于2025年11月27日发表在国际顶尖学术期刊《自然—方法学》上。

为了解决这些限制，研究组开发了FX-Cell及其衍生物，FXcryo-Cell和cryoFX-Cell，以实现难以消化和冷冻保存的植物样品的单细胞RNA测序。通过优化固定缓冲液和最大限度地减少RNA降解，他们的方法确保在高温下有效的细胞壁消化，同时保持高质量的单细胞，即使在-80°C的长期储存后也是如此，并避免使用细胞核，因为细胞核并不代表可翻译信使RNA库。该课题组人员成功构建了水稻分蘖节、野生水稻根茎和大田条件下生长的玉米冠根的高质量细胞图谱。

此外，这些方法能够在单细胞分辨率下精确重建植物急性损伤反应。总的来说，这些进步扩大了植物单细胞基因组学在更广泛物种和组织中的适用性，为植物物种的综合植物细胞地图集铺平了道路。

据了解，植物单细胞RNA测序需要分离高质量的原生质体——没有细胞壁的细胞。然而，许多植物组织和器官对酶消化具有抗性，这对推进植物单细胞多组学研究构成了重大障碍。此外，对于田间种植的作物，缺乏即时的实验室设施是及时制备原生质体的另一个主要挑战。

附：英文原文

Title: FX-Cell: a method for single-cell RNA sequencing on difficult-to-digest and cryopreserved plant samples

Author: Ming, Xin, Wan, Mu-Chun, Zhang, Zheng-Da, Xue, Hao-Chen, Wu, Ya-Qi, Xu, Zhou-Geng, Lian, Heng, Yuan, Meng-Ting, Mai, Yan-Xia, Hu, Ying-Xiong, Liu, Ke, Gao, Jian, Shao, Qiao-Lin, Marchant, D. Blaine, Nelms, Brad, Walbot, Virginia, Wang, Jia-Wei

Issue&Volume: 2025-11-27

Abstract: Single-cell RNA sequencing in plants requires the isolation of high-quality protoplasts—cells devoid of cell walls. However, many plant tissues and organs are resistant to enzymatic digestion, posing a significant barrier to advancing single-cell multi-omics in plant research. Furthermore, for field-grown crops, the lack of immediate laboratory facilities presents another major challenge for timely protoplast preparation. Here, to address these limitations, we developed FX-Cell and its derivatives, FXcryo-Cell and cryoFX-Cell, to enable single-cell RNA sequencing with both difficult-to-digest and cryopreserved plant samples. By optimizing the fixation buffer and minimizing RNA degradation, our approach ensures efficient cell wall digestion at high temperatures while maintaining high-quality single cells, even after long-term storage at 80°C, and circumvents use of nuclei, which are not representative of the pool of translatable messenger RNAs. We successfully constructed high-quality cell atlases for rice tiller nodes, rhizomes of wild rice and maize crown roots grown under field conditions. Moreover, these methods enable the accurate reconstruction of plant acute wounding responses at single-cell resolution. Collectively, these advancements expand the applicability of plant single-cell genomics across a wider range of species and tissues, paving the way for comprehensive Plant Cell Atlases for plant species.

DOI: 10.1038/s41592-025-02900-2

Source: https://www.nature.com/articles/s41592-025-02900-2