Next-generation MOWChIP-seq for high-throughput epigenomic profiling using clinically relevant samples

项目摘要 表观基因组景观中的异常模式与癌症密切相关。 组蛋白修饰的失调沉默肿瘤抑制基因，触发遗传 通过异常DNA修复/复制改变，并导致肿瘤异质性。染色质 免疫沉淀（ChIP）试验是检查表观遗传学的首选技术 体内机制，如组蛋白修饰。然而，该技术在其传统的 形式受到严重的限制。目前流行的ChIP-seq方案需要107个细胞/ 测试，涉及大量的手动处理样品，并需要3 - 4天的时间来生成一个 测序文库大样本量的要求实际上阻止了使用 ChIP-seq用于动物模型和患者等稀缺来源的样本。我们最近 展示了微流体技术（基于微流体振荡洗涤的ChIP-seq或 MOWChIP-seq）用于使用少至100个细胞来分析全基因组蛋白修饰。在 在这个R33项目中，我们将通过促进下一代MOWChIP-seq技术的开发， 高吞吐量，提高集成水平，并通过以下方式进一步验证技术 测试小鼠和人类样本。开发将充分验证MOWChIP的使用- seq用于临床研究中大量样本的高通量表观基因组分析 设置.

Project Summary Aberrant patterns in the epigenomic landscape have been closely associated with cancer. Deregulation of histone modifications silences tumor-suppressor genes, triggers genetic changes via aberrant DNA repair/replication, and contributes to tumor heterogeneity. Chromatin immunoprecipitation (ChIP) assay is the technique of choice for examining epigenetic mechanisms in vivo such as histone modifications. However, the technique in its conventional form suffers from serious limitations. The prevailing ChIP-seq protocol requires 107 cells per test, involves extensive manual handling of the samples, and takes 3-4 days to generate a sequencing library. The requirement of a large sample amount practically prevents the use of ChIP-seq on samples from scarce sources such as animal models and patients. We recently demonstrated a microfluidic technology (microfluidic-oscillatory-washing-based ChIP-seq or MOWChIP-seq) for profiling genome-wide histone modifications using as few as 100 cells. In this R33 project, we will develop the next-generation MOWChIP-seq technology by facilitating high throughput, improving the level of integration, and further validating the technology via testing mouse and human samples. The development will fully validate the use of MOWChIP- seq for high-throughput epigenomic profiling of a large number of samples in clinical research settings.