Building a robust chromatin immunoprecipitation (ChIP) method with substantially improved efficiency

编号 040024004

推送时间 20200525

研究领域 森林培育 

年份 2020 

类型 期刊 

语种 英语

标题 Building a robust chromatin immunoprecipitation (ChIP) method with substantially improved efficiency

来源期刊 Plant Physiology

期 第240期

发表时间 20200423

关键词 Arabidopsis thaliana;  Betula platyphylla;  centrifugal filter;  chromatin immunoprecipitation;  ChIP;  cross-linking reversal;  sucrose; 

摘要 Chromatin immunoprecipitation (ChIP) is the gold-standard method to detect the interactions between proteins and chromatin and is a powerful tool to identify epigenetic modifications. Although ChIP protocols for plant species have been developed, many specific features of plants, especially woody plants, still hinder the efficiency of immunoprecipitation, resulting in inefficient ChIP enrichment and an active demand for a highly efficient ChIP protocol. In the present study using birch (Betula platyphylla) and Arabidopsis (Arabidopsis thaliana) as the research materials, we identified five factors closely associated with ChIP efficiency, including crosslinking, concentrating chromatin using centrifugal filters, using a different immunoprecipitation buffer, rescuing DNA with proteinase K, and using sucrose to increase immunoprecipitation efficiency. Optimization of any these factors can significantly improve ChIP efficiency. Considering these factors together, we developed a robust ChIP protocol which achieved a 14-fold improvement in ChIP enrichment for birch and a more than 6-fold improvement for Arabidopsis compared to the standard ChIP method. As this ChIP method works well in both birch and Arabidopsis, it should also be suitable for other woody and herbaceous species. In addition, this ChIP method enables detection of low-abundance transcription factor-DNA interactions and may extend the application of ChIP in the plant kingdom.

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