脂肪组分染色质免疫共沉淀研究方法的建立

Optimization of chromatin immunoprecipitation for adipose cellular fractions

目的:优化肾周等部位脂肪组织各组分染色质免疫共沉淀(chromatin immunoprecipitation,ChIP)的实验方案。方法:测试了脂肪各组分细胞分离和染色质提取的不同方法,以优化脂肪组分的ChIP实验方法。结果:通过优化ChIP实验各步骤,减少了细胞损失、防止油脂污染,从而提高了脂肪各组分染色质提取的质量和数量。从4只小鼠的5个脂肪部位成功获得ChIP实验所需的染色质,为成熟脂肪细胞和血管基质组分(stromal vascular fraction,SVF)细胞生成了高质量的组蛋白修饰图谱。结论:本研究通过优化适合脂肪组织的ChIP实验方法,提高了新鲜脂肪组分染色质的数量和质量,可为成熟脂肪细胞和SVF细胞生成高质量组蛋白修饰图谱。

Objective:To explore the optimization of chromatin preparation from perirenal and other adipose cellular fractions for chromatin immunoprecipitation(ChIP)-seq.Methodology:Different ways for cell isolation and chromatin extraction to optimize the ChIP protocol on adipose fractions were tested.Results:With our optimized protocol,the cell loss could be reduced,lipid contamination could be prevented and the quality and quantity of chromatin from mass-limited adipose cellular fractions could be improved.Enough chromatin required for the ChIP experiment for each adipose depot from 4 mice,and resulted in high performance histone modification map for mature adipocytes and stromal vascular fraction(SVF)cells could be obtained.Conclusion:We described an optimized ChIP protocol that improved the quantity and quality of chromatin from fresh adipose fractions and resulted in high performance histone modification map for mature adipocytes and SVF cells.