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引用次数: 0
摘要
染色质免疫沉淀(ChIP)是一种特征明确的程序,用于揭示蛋白质-DNA 相互作用的特定模式,并确定蛋白质在 DNA 上的结合位点。ChIP 已被用于研究果蝇生物学的许多方面,包括神经生物学。本方案详细介绍了如何从果蝇触角和大脑中制备交联染色质,然后进行免疫沉淀(X-ChIP)。我们首先介绍了组织解剖、用甲醛进行染色质交联、交联淬灭、组织匀浆和超声剪切染色质。此外,我们还以果蝇大脑样本为例,介绍了如何优化超声效率、固定时间和浓度。这些参数对 ChIP 的成功至关重要。
Sample Preparation for Chromatin Immunoprecipitation (ChIP) from Drosophila Antennal and Brain Samples.
Chromatin immunoprecipitation (ChIP) is a well-characterized procedure used to reveal specific patterns of protein-DNA interactions and identify the binding sites of proteins on DNA. ChIP has been used to study many aspects of Drosophila biology, including neurobiology. This protocol describes in detail how to prepare cross-linked chromatin from Drosophila antennae and brains followed by immunoprecipitation (X-ChIP). We first describe tissue dissection, chromatin cross-linking with formaldehyde, quenching of the cross-linking, homogenization of tissues, and sonication for shearing the chromatin. Additionally, we describe how to optimize the sonication efficiency and fixation time and concentration using Drosophila brain samples as an example. These parameters are crucial for successful ChIP.
Cold Spring Harbor protocolsBiochemistry, Genetics and Molecular Biology-Biochemistry, Genetics and Molecular Biology (all)
CiteScore
3.00
自引率
0.00%
发文量
163
期刊介绍:
Cold Spring Harbor Laboratory is renowned for its teaching of biomedical research techniques. For decades, participants in its celebrated, hands-on courses and users of its laboratory manuals have gained access to the most authoritative and reliable methods in molecular and cellular biology. Now that access has moved online. Cold Spring Harbor Protocols is an interdisciplinary journal providing a definitive source of research methods in cell, developmental and molecular biology, genetics, bioinformatics, protein science, computational biology, immunology, neuroscience and imaging. Each monthly issue details multiple essential methods—a mix of cutting-edge and well-established techniques.