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An efficient immunodetection method for histone modifications in plants

机译:一种有效的植物组蛋白修饰免疫检测方法

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Background Epigenetic mechanisms can be highly dynamic, but the cross-talk among them and with the genome is still poorly understood. Many of these mechanisms work at different places in the cell and at different times of organism development. Covalent histone modifications are one of the most complex and studied epigenetic mechanisms involved in cellular reprogramming and development in plants. Therefore, the knowledge of the spatial distribution of histone methylation in different tissues is important to understand their behavior on specific cells. Results Based on the importance of epigenetic marks for biology, we present a simplified, inexpensive and efficient protocol for in situ immunolocalization on different tissues such as flowers, buds, callus, somatic embryo and meristematic tissue from several plants of agronomical and biological importance. Here, we fully describe all the steps to perform the localization of histone modifications. Using this method, we were able to visualize the distribution of H3K4me3 and H3K9me2 without loss of histological integrity of tissues from several plants, including Agave tequilana, Capsicum chinense, Coffea canephora and Cedrela odorata, as well as Arabidopsis thaliana. Conclusions There are many protocols to study chromatin modifications; however, most of them are expensive, difficult and require sophisticated equipment. Here, we provide an efficient protocol for in situ localization of histone methylation that dispenses with the use of expensive and sensitive enzymes. The present method can be used to investigate the cellular distribution and localization of a wide array of proteins, which could help to clarify the biological role that they play at specific times and places in different tissues of various plant species.
机译:背景表观遗传机制可能是高度动态的,但是它们之间以及与基因组的串扰仍然知之甚少。这些机制中的许多机制在细胞的不同位置和生物体发育的不同时间起作用。共价组蛋白修饰是涉及植物细胞重编程和发育的最复杂且研究最深入的表观遗传机制之一。因此,了解组蛋白甲基化在不同组织中的空间分布对于了解它们在特定细胞上的行为非常重要。结果基于表观遗传标记对生物学的重要性,我们提出了一种简化,廉价且有效的协议,用于在几种具有农艺和生物学重要性的植物上的不同组织(例如花朵,芽,愈伤组织,体细胞胚和分生组织)上进行原位免疫定位。在这里,我们充分描述了执行组蛋白修饰本地化的所有步骤。使用这种方法,我们能够可视化H3K4me3和H3K9me2的分布,而不会损失几种植物的组织的组织学完整性,这些植物包括龙舌兰龙舌兰,辣椒,香叶咖啡和雪松以及拟南芥。结论有很多研究染色质修饰的方法。然而,它们大多数昂贵,困难并且需要复杂的设备。在这里,我们为组蛋白甲基化的原位定位提供了一种有效的方案,而无需使用昂贵且敏感的酶。本方法可用于研究多种蛋白质的细胞分布和定位,这可有助于阐明它们在特定时间和位置在各种植物物种的不同组织中发挥的生物学作用。

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