Salinas Edward A, Edwards Yvonne J K
Department of Biochemistry and Molecular Genetics, Heersink School of Medicine, University of Alabama at Birmingham, Birmingham, AL, USA.
Gregory Fleming James Cystic Fibrosis Research Center, Heersink School of Medicine, University of Alabama at Birmingham, Birmingham, AL, USA.
Adv Exp Med Biol. 2025;1485:75-95. doi: 10.1007/978-981-96-9428-0_6.
Circular RNAs (circRNAs) play various roles in gene regulation and are known to be associated with the development of disease. They can act as microRNA sponges, interact with RNA-binding proteins, and even contain open reading frames (ORFs) that are translated to generate proteins. Scientists hope to better understand these covalently closed ring-like structures as they are not well-characterized.Here, we present CircRNAFlow, a free and open-source Nextflow-based pipeline for in silico circRNA identification and characterization. This next-generation sequencing (NGS) pipeline takes paired-end FASTQ files as input and produces tables and graphical images that report identified circRNAs and their corresponding characterizations. Reports include CircTest to compare circRNA expression to host gene expression and across groups. Characterization includes ClusterProfiler, CRAFT (CircRNA Function prediction Tool), and deepTarget, which explore the regulatory roles of identified circRNAs.In this chapter, we discuss the bioinformatics analyses carried out by the incorporated third-party software of CircRNAFlow [namely, FastQC, Flexbar, Bowtie2, STAR (Spliced Transcripts Alignment to a Reference), DCC (detect circRNAs from chimeric reads), CircTest, CRAFT, ClusterProfiler, and deepTarget]. This chapter, along with a companion GitHub repository, provides a tutorial on CircRNAFlow, including installation, running the software, and interpretation of the outputs. This chapter also provides some troubleshooting tips.
环状RNA（circRNAs）在基因调控中发挥着多种作用，并且已知与疾病的发展相关。它们可以充当微小RNA海绵，与RNA结合蛋白相互作用，甚至包含可翻译产生蛋白质的开放阅读框（ORF）。由于这些共价闭合的环状结构尚未得到充分表征，科学家们希望能更好地了解它们。在此，我们介绍CircRNAFlow，这是一个基于Nextflow的免费开源流程，用于在计算机上鉴定和表征环状RNA。这个新一代测序（NGS）流程以双端FASTQ文件作为输入，并生成报告已鉴定的环状RNA及其相应特征的表格和图形图像。报告包括CircTest，用于比较环状RNA与宿主基因的表达以及不同组之间的表达。特征分析包括ClusterProfiler、CRAFT（环状RNA功能预测工具）和deepTarget，它们用于探索已鉴定的环状RNA的调控作用。在本章中，我们讨论了由CircRNAFlow整合的第三方软件（即FastQC、Flexbar、Bowtie2、STAR（与参考序列比对的剪接转录本）、DCC（从嵌合读段中检测环状RNA）、CircTest、CRAFT、ClusterProfiler和deepTarget）所进行的生物信息学分析。本章与配套的GitHub仓库一起提供了关于CircRNAFlow的教程，包括软件安装、运行以及输出结果的解读。本章还提供了一些故障排除提示。
