干化对粗枝云杉体胚解剖结构及细胞壁重塑基因表达的影响

Effects of Desiccation on Anatomical Structure of Somatic Embryo and Expression of Cell Wall Remodelin g Gene of Picea asperata

[目的]探究干化对云杉体胚解剖结构的影响,挖掘导致解剖结构改变的关键基因,初步明确干化促进体胚萌发的原因。[方法]以高胚性粗枝云杉细胞系为研究材料诱导体胚发生,干化处理不同时间后进行体胚表型观察及萌发率的测定。利用超景深显微镜、扫描电镜和透射电镜从不同层次观察干化体胚与未干化体胚的组织和细胞结构差异,并结合转录组测序分析干化前后体胚的基因表达变化。[结果]干化显著改变了粗枝云杉体胚形态,干化后体胚胚根呈红色,子叶呈绿色。显微观察发现干化后体胚的细胞呈现皱缩且分布不规则的状态,高倍视野下细胞壁不再具有刚性,表现出松弛扭曲状。转录组分析发现参与细胞壁重塑和影响细胞壁机械性能的多个XTHs、EXPAs和PMEs转录本在干化和未干化体胚中差异表达,且多上调表达,与显微观察发现干化体胚细胞壁形态变化的结果相符,这几类基因可能协同调控该过程,影响体胚萌发。[结论]干化过程中细胞壁重塑基因差异表达导致体胚细胞壁松弛,刚性结构消失,解除细胞分化和生长的限制,是干化促进体胚萌发的潜在原因。

[Objective]To explore the effects of desiccation on the anatomical structure of somatic embryo of Picea asperata,identify the key genes that lead to the changes in anatomical structure,and preliminar-ily explore the reasons why desiccation promotes somatic embryo germination.[Method]The high em-bryonic cell line of Picea asperata was used to induce somatic embryogenesis.The phenotypic observa-tion and germination rate of somatic embryos were determined after desiccation treatment for different time.Ultra depth of field microscope,scanning electron microscope and transmission electron microscope were used to observe the differences of tissue and cell structure and morphology between desiccated so-matic embryos and non-desiccated somatic embryos,and transcriptome sequencing was combined to analyze the changes of gene expression between desiccated and non-desiccated somatic embryos.[Results]The somatic embryo morphology of spruce was significantly changed after desiccation.The rad-icle turned red and the cotyledon turned green after desiccation.Microscopic observation showed that the cells of the desiccated somatic embryo were shrunken and irregularly distributed.At high magnification,the cell wall was no longer rigid,showing a relaxed and twisted state.Transcriptome analysis found that several transcripts of XTHs,EXPAs and PMEs involved in cell wall remodeling and affecting the mechanic-al properties of cell walls were significantly differentially expressed between desiccated and non-desic-cated somatic embryos,and their expressions were mostly up-regulated,which was consistent with the results of microscopic observation on the morphological changes of desiccated somatic embryos.These genes may synergically regulate the process of desiccation and affect the germination of somatic embryos.[Conclusion]The differential expression of cell wall remodeling genes during desiccation leads to the relaxa-tion of somatic embryo cell wall,the disappearance of rigid structure,and the release of cell differentiation and growth restrictions,which is a potential reason for desiccation to promote somatic embryo germination.