State-of-the-Art Circular RNA Analytics Using the Circtools Software Suite

Circular RNAs (circRNAs) are types of RNA molecules that have been discovered relatively recently and have been found to be widely expressed in eukaryotic cells. Unlike canonical linear RNA molecules, circRNAs form a covalently closed continuous loop structure without a 5′ or 3′ end. They are generated by a process called back-splicing, in which a downstream splice donor site is joined to an upstream splice acceptor site. CircRNAs have been found to play important roles in various biological processes, including gene regulation, alternative splicing, and protein translation. They can act as sponges for microRNAs or RNA-binding proteins and can also encode peptides or proteins. Additionally, circRNAs have been implicated in several diseases, including cancer, neurological disorders, and cardiovascular diseases. This protocol provides all necessary steps to detect and analyze circRNAs in silico from RNA sequencing data using the circtools circRNA analytics software suite. The protocol starts from raw sequencing data with circRNA detection via back-splice events and includes statistical testing of circRNAs as well as primer design for follow-up wet lab experiments.