美国华盛顿大学Jay Shendure课题组和美国加州大学旧金山分校Nadav Ahituv课题组合作开发出了系统评价大规模并行报告基因分析的方法，并对9种MPRA设计进行了比较。该研究于2020年10月12日发表于《自然—方法学》。

该研究团队使用9种不同MPRA设计，筛选了用于HepG2调控活动的包含2440种候选肝增强子和对照的库。该团队确定了与表观遗传和序列水平特征相关的细微但重要的差异，以及动态范围和再现性的差异。课题组人员还验证了，至少对于它们的库和设计而言，增强子的活动在很大程度上与方向无关。

最后，该研究团队组装和测试相同的增强剂192-mers，354-mers以及678-mers，观察到相当大的差异。这项工作提供了一个框架，用于高通量报告基因分析的实验设计，表明测试元素序列内容的拓展，并在较小程度的精确测定，会影响MPRA结果。

据了解，大规模并行报告基因分析(MPRAs)功能性地并行筛选上千用于调控活动的基因序列。到目前为止，仅有非常有限的研究系统性比较了不同MPRA设计。

附：英文原文

Title: A systematic evaluation of the design and context dependencies of massively parallel reporter assays

Author: Jason C. Klein, Vikram Agarwal, Fumitaka Inoue, Aidan Keith, Beth Martin, Martin Kircher, Nadav Ahituv, Jay Shendure

Issue&Volume: 2020-10-12

Abstract: Massively parallel reporter assays (MPRAs) functionally screen thousands of sequences for regulatory activity in parallel. To date, there are limited studies that systematically compare differences in MPRA design. Here, we screen a library of 2,440 candidate liver enhancers and controls for regulatory activity in HepG2 cells using nine different MPRA designs. We identify subtle but significant differences that correlate with epigenetic and sequence-level features, as well as differences in dynamic range and reproducibility. We also validate that enhancer activity is largely independent of orientation, at least for our library and designs. Finally, we assemble and test the same enhancers as 192-mers, 354-mers and 678-mers and observe sizable differences. This work provides a framework for the experimental design of high-throughput reporter assays, suggesting that the extended sequence context of tested elements and to a lesser degree the precise assay, influence MPRA results. 

DOI: 10.1038/s41592-020-0965-y

Source: https://www.nature.com/articles/s41592-020-0965-y