Understanding how regulatory sequences control gene expression is fundamental to explain how phenotypes arise in health and disease. Traditional reporter assays inform about function of individual regulatory elements, typically in isolation. However, regulatory elements must ultimately be understood by perturbing them within their genomic environment and developmental- or tissue-specific contexts. This is technically challenging; therefore, few regulatory elements have been characterized in vivo. Here, we used inducible Cas9 and multiplexed guide RNAs to create hundreds of mutations in enhancers/promoters and 3′ UTRs of 16 genes in C. elegans. To quantify the consequences of mutations on expression, we developed a targeted RNA sequencing strategy across hundreds of mutant animals. We were also able to systematically and quantitatively assign fitness cost to mutations. Finally, we identified and characterized sequence elements that strongly regulate phenotypic traits. Our approach enables highly parallelized, functional analysis of regulatory sequences in vivo.

中文翻译：

---

体内调节序列的平行遗传

了解调节序列如何控制基因表达对于解释表型如何在健康和疾病中产生至关重要。传统的报告基因检测通常会单独地告知单个调节元件的功能。但是，必须通过在其基因组环境和特定于发育或组织的环境中对其进行干扰来最终理解其调控元件。这在技术上具有挑战性；因此，体内几乎没有调节元件的特征。在这里，我们使用了可诱导的Cas9和多重引导RNA，在秀丽隐杆线虫的16个基因的增强子/启动子和3'UTR中创建了数百个突变。为了量化突变对表达的影响，我们开发了针对数百种突变动物的靶向RNA测序策略。我们还能够系统地和定量地将适应性成本分配给突变。最后，我们鉴定并表征了强烈调节表型性状的序列元件。我们的方法可以在体内对调节序列进行高度并行的功能分析。