Accurate Reconstruction of Circular RNAs from Complex Rolling Circular Long Reads with CircPlex

Area of Science:

• Molecular Biology
• Genomics
• Bioinformatics

Background:

• Circular RNAs (circRNAs) are crucial regulatory molecules.
• Long-read sequencing with Rolling Circle Amplification (RCA) is a key method for detecting circRNAs.
• Standard analysis of RCA reads can be confounded by complex repeat patterns.

Purpose of the Study:

• To develop a novel computational approach to accurately identify authentic circRNA sequences from RCA long reads.
• To address the challenge of complex repeat patterns in RCA data that can lead to misidentification of circRNAs.
• To improve the sensitivity and comprehensiveness of circRNA detection.

Main Methods:

• Development of CircPlex algorithm for extracting true circRNA sequences from complex repeat units.
• Analysis of long reads generated by RCA and long-read sequencing.
• Comparison of CircPlex results with existing annotation tools (isoCirc) and circRNA databases.

Main Results:

• Identified a significant fraction of long reads with complex repeat patterns, including reverse complements.
• CircPlex successfully extracted authentic circRNA sequences, overcoming limitations of standard consensus prediction.
• Recovered a substantial number of previously ignored back-splice junctions (BSJs) and full-length circRNA sequences.

Conclusions:

• CircPlex enhances the detection of circRNAs by accurately interpreting complex read patterns.
• Leveraging partially repetitive reads from RCA-based sequencing significantly increases circRNA discovery.
• This approach provides a more comprehensive view of the circular transcriptome, uncovering novel isoforms.