Developing a standardized kit and normal reference profile for circular RNAs in the nervous system

开发神经系统中环状 RNA 的标准化试剂盒和正常参考谱

• 批准号: 8841991
• 负责人: Jarret Glasscock
• 金额: $ 16.15万
• 依托单位: COFACTOR GENOMICS, LLC
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-01 至 2015-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8841991
• 关键词: Address; Award; Binding; Biological Assay; Biological Markers; Brain; Brain region; Cell Maintenance; Cerebral cortex; Cognition Disorders; Cytoplasm; Data; Data Set; Development; Disease; Evaluation; Event; Exons; Fostering; Functional RNA; Future; Gene Expression Profile; Genomics; Goals; Harvest; Health; Hippocampus (Brain); Histocompatibility Testing; Human; Human Biology; In Vitro; Libraries; Marketing; Mental disorders; Messenger RNA; Methods; MicroRNAs; Monitor; Nervous system structure; Neuroglia; Neurons; Pathway interactions; Phase; Play; Poly A; Polyadenylation; Population; Post-Transcriptional Regulation; Preparation; Protein Isoforms; Protocols documentation; RNA; RNA amplification; RNA-Directed DNA Polymerase; Relative (related person); Research; Research Contracts; Research Personnel; Resources; Ribonucleases; Ribosomal RNA; Role; Sampling; Scientist; Sequence Analysis; Sequence Homology; Services; Site; Substance abuse problem; Testing; Tissues; Transcript; Translating; Translations; Validation; Work; brain tissue; cell type; circular RNA; cofactor; efficacy testing; experience; gene panel; infancy; insight; interest; nervous system disorder; neurodevelopment; neuron development; next generation sequencing; novel; phase 2 study; protocol development; public health relevance; tool

DESCRIPTION (provided by applicant): Modified ribonucleic acids (RNA) are important regulators of translation and other cellular pathways. Non- coding RNAs like microRNAs play significant roles in neuronal development, and can serve as biomarkers for psychiatric disorders. Another such modified RNA is circular RNA (circRNA), a non-coding transcript present in the cytoplasm. Since circRNAs are present in lower abundance than other RNA molecules, and share sequence homology with mRNA, they are difficult to isolate from total RNA. To facilitate identification and examination of these molecules, we propose to develop a streamlined approach that uses selective reduction of linear and ribosomal RNAs combined with a novel reverse transcriptase to enrich for circular RNAs. To optimize for robust reduction and amplification, we will test a range of conditions and assess their relative efficiencies using qRT-PCR specific for known circRNAs and synthetic spiked-in linear RNAs. We will use our validated approach to generate circRNA profiles of multiple brain tissues, which will be of significant benefit to researchers interested in understanding the role of circRNA in neuronal development, substance abuse, and psychiatric disorders. As there is no kit on the market which addresses the need for easy circRNA enrichment, development of this protocol will foster broader research in this nascent field and further understanding of the role of circRNAs.

描述(申请人提供)：修饰的核糖核酸(RNA)是翻译和其他细胞途径的重要调节因子。像microRNAs这样的非编码RNA在神经元发育中发挥着重要作用，可以作为精神疾病的生物标记物。另一种这种修饰的RNA是环状RNA(CircRNA)，它是一种存在于细胞质中的非编码转录本。由于CircRNAs的丰度低于其他RNA分子，并且与mRNA具有序列同源性，因此很难从总RNA中分离出它们。为了便于鉴定和检测这些分子，我们建议开发一种简化的方法，使用选择性地还原线性和核糖体RNA，并结合一种新的逆转录酶来丰富环状RNA。为了优化稳健的还原和扩增，我们将测试一系列条件，并使用针对已知CircRNA和合成掺入线性RNA的qRT-PCR来评估它们的相对效率。我们将使用我们经过验证的方法来生成多个脑组织的CircRNA图谱，这将对有兴趣了解CircRNA在神经元发育、物质滥用和精神障碍中的作用的研究人员有重大好处。由于市场上还没有一种试剂盒可以满足对简单CircRNA浓缩的需求，该方案的开发将促进这一新兴领域的更广泛研究，并进一步了解CircRNA的作用。