Key message

We present a detailed protocol for isolation of single sperm cells and transcriptome analysis to study variation in gene expression between sperm cells.

Abstract

Male gametophyte development in flowering plants begins with a microspore mother cell, which upon two consecutive cell divisions forms a mature pollen grain containing a vegetative nucleus and two sperm cells. Pollen development is a highly dynamic process, involving changes at both the transcriptome and epigenome levels of vegetative nuclei and the pair of sperm cells that have their own cytoplasm and nucleus. While the overall transcriptome of Arabidopsis pollen development is well documented, studies at single-cell level, in particular of sperm cells, are still lacking. Such studies would be essential to understand whether and how the two sperm cells are transcriptionally different, in particular once the pollen tube grows through the transmitting tissue of the pistil. Here we describe a detailed protocol for isolation of single sperm cells from growing pollen tubes and analysis of their transcriptome.

中文翻译：

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单细胞分辨率下拟南芥精子的转录组学。

关键信息

我们提出了用于分离单个精子细胞和转录组分析的详细协议，以研究精子细胞之间基因表达的差异。

抽象

开花植物中雄配子体的发育始于小孢子母细胞，该小孢子母细胞在两次连续的细胞分裂后形成成熟的花粉粒，其中含有营养核和两个精子细胞。花粉发育是一个高度动态的过程，涉及营养细胞核以及具有其自身细胞质和细胞核的一对精子细胞的转录组和表观基因组水平的变化。尽管拟南芥花粉发育的总体转录组已有充分文献记载，但仍缺乏单细胞水平，尤其是精子细胞水平的研究。这样的研究对于理解两个精子细胞是否以及如何转录是至关重要的，特别是一旦花粉管通过雌蕊的传播组织生长时。