Empirical Detection of Small RNA Targets

在包括植物和动物在内的许多多细胞生物中，自然产生的21至24个核苷酸大小的小rna是基因表达的普遍特征。这些小RNA最终作为特异性决定因素，根据小RNA和靶标之间的沃森-克里克碱基配对，指导抑制蛋白复合物靶向信使RNA。因此，与经典的蛋白质编码基因不同，小rna的表达并不直接影响表型。只有通过研究小rna的靶标，才能了解它们的功能。已经描述了几种预测小rna靶标的计算方法，并且有一些实验方法可用于每次测试一个预测。然而，目前还没有能够对小RNA靶点进行无偏、全基因组和经验性检测的方法。该项目旨在开发和实施技术，将首次在实验中识别体内所有小RNA靶标。所有生物的所有特征最终都是由基因控制的。并不是所有的基因都是一直“开着”的——对单个基因“开-关”状态的精细调节是生命的绝对必要。在动植物等复杂生物体中，所谓的“小rna”在调节特定“靶”基因方面发挥着非常重要的作用。因此，确定这些小rna的靶标对于理解基因调控至关重要。该项目将开发第一个用于鉴定小RNA靶标的实验性、无偏见的方法。本研究将广泛影响小rna介导的基因调控的研究，也将有助于下一代科学家的跨学科培训。

Naturally occurring small RNAs of 21 to 24 nucleotides in size are a ubiquitous feature of gene expression in many multicellular organisms, including plants and animals. These small RNAs ultimately function as specificity determinants that guide repressive protein complexes to target messenger RNAs based upon Watson-Crick base-pairing between the small RNA and target. Therefore, unlike classical protein-coding genes, expression of small RNAs does not directly influence phenotype. It is only though study the targets of small RNAs that their functions may be understood. Several computational methods have been described that predict the targets of small RNAs, and there are some experimental methodologies available for testing these predictions on a one at a time basis. However, there is currently no methodology capable of unbiased, genome-wide, and empirical detection of small RNA targets. This project seeks to develop and implement techniques that will, for the first time, experimentally identify all small RNA targets in vivo. All characteristics of every living thing are ultimately controlled by genes. Not all genes are "on" all of the time - fine regulation of the "on-off" states of individual genes is an absolute necessity for life. So-called "small RNAs" play a very important role in regulating specific "target" genes in complex organisms like plants and animals. Identifying the targets of these small RNAs is thus critical to understanding gene regulation. This project will develop the first experimental, non-biased methodology for the identification of small RNA targets. This research will broadly impact the study of small RNA-mediated gene regulation, and will also contribute to the interdisciplinary training of the next generation of scientists.