Native Chromatin Immunoprecipitation Using Murine Brain Tumor Neurospheres.
Native Chromatin Immunoprecipitation Using Murine Brain Tumor Neurospheres.
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DOI：
10.3791/57016
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发表时间：
2018-01-29
期刊：
影响因子：
--
通讯作者：
Castro MG
中科院分区：
文献类型：
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作者：
Mendez FM;Núñez FJ;Zorrilla-Veloz RI;Lowenstein PR;Castro MG
表观遗传修饰可能参与了胶质瘤的发生和发展。癌基因和抑癌基因启动子和调节区的甲基化和乙酰化的改变可导致基因表达的变化，在脑肿瘤的发病机制中发挥重要作用。天然染色质免疫沉淀(CHIP)是一种流行的技术，它可以检测修饰或其他与DNA紧密结合的蛋白质。与交联型芯片不同的是，在天然芯片中，细胞不会被甲醛处理以共价连接蛋白质和DNA。这是有利的，因为有时交联会固定只与DNA短暂相互作用的蛋白质，而在基因调控中没有功能意义。此外，抗体通常是针对未固定的多肽而产生的。因此，在国产芯片中提高了抗体的特异性。然而，重要的是要记住，天然芯片只适用于研究组蛋白或其他与DNA紧密结合的蛋白质。本方案描述了小鼠脑肿瘤神经球上的天然染色质免疫沉淀。
Epigenetic modifications may be involved in the development and progression of glioma. Changes in methylation and acetylation of promoters and regulatory regions of oncogenes and tumor suppressors can lead to changes in gene expression and play an important role in the pathogenesis of brain tumors. Native chromatin immunoprecipitation (ChIP) is a popular technique that allows the detection of modifications or other proteins tightly bound to DNA. In contrast to cross-linked ChIP, in native ChIP, cells are not treated with formaldehyde to covalently link protein to DNA. This is advantageous because sometimes crosslinking may fix proteins that only transiently interact with DNA and do not have functional significance in gene regulation. In addition, antibodies are generally raised against unfixed peptides. Therefore, antibody specificity is increased in native ChIP. However, it is important to keep in mind that native ChIP is only applicable to study histones or other proteins that bind tightly to DNA. This protocol describes the native chromatin immunoprecipitation on murine brain tumor neurospheres.