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染色质免疫沉淀法筛选转录因子Nanog调控的目的基因

Identification of the Downstream Targets of Nanog by Chromatin Immunoprecipitation
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摘要 目的:利用染色质免疫沉淀法筛选转录因子Nanog调控的目的基因。方法:用甲醛固定胚胎干细胞,利用超声将其染色质随机断裂成0.5～1.0kb的染色质片段,通过免疫沉淀富集与Nanog结合的DNA片段,回复交联后分离纯化DNA片段,最终用凝胶迁移阻滞实验验证Nanog与DNA片段的结合。结果:我们发现fzr为Nanog调控的目的基因,并通过电泳迁移率变动分析证明了二者的结合。结论:Fzr在细胞周期调控中发挥重要作用,这为阐明Nanog的作用机理奠定基础。 Objective： To identify the downstream targets of Nanog by chromatin immunoprecipitation. Methods： By formaldehyde cross-linking and sonicationg, the chromatin of embryonic stem cells was splitted to fragments with an aver- age length approximately 0.5-1 kb. The DNA fragments binding to targeted protein Nanog were riched by immunoprecipi- tation, and were isolatede and purificated after reverse crosslinks. The combination of Nanog and DNA fragments was proved by electrophoretic mobility shift assay（EMSA）. Results： Our data shows that the following screened gene is fzr. We proved the results by EMSA. Conclusion： Fzr is important for the regulation of cell cycle. This elucidates the mechanism of Nanog function elementarily.
作者 林艳丽 周艳荣 阎新龙 熊福银 田利源 吴晓洁 卢建申 邓继先 陈红星
机构地区 军事医学科学院生物工程研究所
出处 《生物技术通讯》 CAS 2009年第6期776-779,共4页 Letters in Biotechnology
基金 国家自然科学基金(30570916)
关键词 NANOG 染色质免疫沉淀 fzr基因 电泳迁移率变动分析 Nanog chromatin immunoprecipitation fzr gene electrophoretic mobility shift assay
分类号 Q75 [生物学—分子生物学] Q503 [生物学—生物化学]
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参考文献17

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生物技术通讯
2009年 第6期

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