Pollen represents the male sexual lineage in flowering plants. At maturity, pollen grains are composed of a companion vegetative cell with embedded sperm. During pollen development, these two cell types acquire vastly differing cell fates. Underlying this differential fate acquisition is dramatic reconfiguration of pollen chromatin that is highly evident at a cytological level. The precise link between histone mark deposition and fate acquisition remains largely unexplored, which in part has been hindered by the difficulty in working with pollen in model plant species like Arabidopsis. Here, we describe a simple and robust protocol to isolate Arabidopsis pollen nuclei and immunostain for histone marks. Plant growth aside, the protocol can be performed over 2 days with few Arabidopsis plants, thus allowing multiple genotypes to be analysed in parallel. We also describe a method to de-mask epitopes through antigen retrieval, which vastly improves the signal for antibodies that target heterochromatic histone marks.

中文翻译：

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用于免疫染色拟南芥花粉核的简单而可靠的协议。

花粉代表开花植物中的男性性世系。成熟时，花粉粒由伴生的营养细胞和精子嵌入组成。在花粉发育过程中，这两种细胞类型具有截然不同的细胞命运。这种差异性命运的获得基础是花粉染色质的戏剧性重构，这在细胞学水平上非常明显。组蛋白标记沉积与命运获得之间的确切联系仍未得到充分探索，部分原因是难以在拟南芥等模式植物物种中使用花粉处理。在这里，我们描述了一个简单而健壮的协议，以隔离拟南芥花粉核和组蛋白标记的免疫染色。除了植物生长，该方案可以在2天的时间内完成，几乎不需要拟南芥植物，因此允许并行分析多个基因型。我们还描述了一种通过抗原检索来掩盖表位的方法，该方法极大地改善了针对异源组蛋白标记的抗体的信号。