Laser microdissection and fluorescence in situ hybridization reveal the tissue-specific gene expression in the ovules of P. tabulaeformis Carr

IF 4 3区生物学 Q1 PLANT SCIENCES

Journal of plant physiology Pub Date : 2025-04-22 DOI:10.1016/j.jplph.2025.154500

Xinyu Zhang , Binli Chen , Xiaoxin Song , Yingqi Wang , Caixia Zheng , Zaixin Gong

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Abstract

Ovules are important carriers for seed plant reproduction, and ovules of gymnosperms are composed mainly of female gametophyte (FG) and adjacent diploid tissue (ADT). To investigate tissue-specific genes in the ovules of Pinus tabulaeformis Carr., we used laser microdissection (LMD) to separate FGs and ADTs, and performed linear amplification to construct cDNA libraries, obtaining a total of 156 expressed sequence tags (EST). Furthermore, some differentially expressed genes between FG and ADT of P. tabulaeformis ovule were screened by the analysis of EST. In addition, the expression levels of key genes in fertile line (FL) and sterile line (SL) ovules during development were verified by RT-qPCR, and we found that both PtRPL7a and PtDHN4 were more highly expressed in FL in each period (at least 1.7 times that of SL). Finally, fluorescence in situ hybridization (FISH) was used to reveal the temporal and spatial expression patterns of PtRPL7a and PtDHN4 in the ovules of P. tabuliformis during ovule development between FL and SL. Our results indicate that the expression levels and the locations of PtRPL7a and PtDHN4 show significant differences in different tissues during ovule development between FL and SL. This study further elucidates the molecular mechanism of the ovule abortion of P. tabulaeformis and provides a theoretical basis for the germplasm optimization of gymnosperms.

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激光显微解剖和荧光原位杂交技术揭示了油棕胚珠中组织特异性基因的表达

胚珠是种子植物繁殖的重要载体，裸子植物的胚珠主要由雌性配子体（FG）和相邻的二倍体组织（ADT）组成。目的：研究油松胚珠的组织特异性基因。利用激光显微解剖（LMD）分离FGs和ADTs，并进行线性扩增构建cDNA文库，共获得156个表达序列标签（EST）。此外，通过EST分析筛选油油树胚珠FG与ADT之间的差异表达基因。此外，通过RT-qPCR验证了关键基因在可育系（FL）和不育系（SL）胚珠发育过程中的表达水平，我们发现PtRPL7a和PtDHN4在FL胚珠各时期的表达水平都更高（至少是SL的1.7倍）。最后,利用荧光原位杂交技术（FISH）揭示了PtRPL7a和PtDHN4在油松胚珠发育过程中的时空表达模式，结果表明，PtRPL7a和PtDHN4在油松胚珠发育过程中不同组织中的表达水平和位置存在显著差异。本研究进一步阐明了油松胚珠败育的分子机制，并为油松胚珠败育的研究提供了科学依据裸子植物种质资源优化的理论基础。

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来源期刊

Journal of plant physiology 生物-植物科学

CiteScore

7.20

自引率

4.70%

发文量

196

审稿时长

32 days

期刊介绍： The Journal of Plant Physiology is a broad-spectrum journal that welcomes high-quality submissions in all major areas of plant physiology, including plant biochemistry, functional biotechnology, computational and synthetic plant biology, growth and development, photosynthesis and respiration, transport and translocation, plant-microbe interactions, biotic and abiotic stress. Studies are welcome at all levels of integration ranging from molecules and cells to organisms and their environments and are expected to use state-of-the-art methodologies. Pure gene expression studies are not within the focus of our journal. To be considered for publication, papers must significantly contribute to the mechanistic understanding of physiological processes, and not be merely descriptive, or confirmatory of previous results. We encourage the submission of papers that explore the physiology of non-model as well as accepted model species and those that bridge basic and applied research. For instance, studies on agricultural plants that show new physiological mechanisms to improve agricultural efficiency are welcome. Studies performed under uncontrolled situations (e.g. field conditions) not providing mechanistic insight will not be considered for publication. The Journal of Plant Physiology publishes several types of articles: Original Research Articles, Reviews, Perspectives Articles, and Short Communications. Reviews and Perspectives will be solicited by the Editors; unsolicited reviews are also welcome but only from authors with a strong track record in the field of the review. Original research papers comprise the majority of published contributions.