A step-by-step guide to analyzing CAGE data using R/Bioconductor

Area of Science:

• Genomics
• Bioinformatics
• Molecular Biology

Background:

• Cap Analysis of Gene Expression (CAGE) is a widely used 5'-end sequencing technique.
• CAGE enables the identification and quantification of Transcription Start Sites (TSSs) and enhancers in a single experiment.

Purpose of the Study:

• To provide a case study demonstrating the use of the CAGEfightR package for orchestrating CAGE data analysis within the Bioconductor project.
• To outline a workflow for comprehensive CAGE data analysis, from basic identification to advanced interaction analysis.

Main Methods:

• Utilizing the CAGEfightR package for analysis of CAGE data, starting from BigWig files.
• Implementing R-code for identifying, quantifying, and annotating TSSs and enhancers.
• Performing advanced analyses including TSS-enhancer pair identification, enhancer clustering, differential expression analysis, and alternative TSS usage.

Main Results:

• The workflow successfully integrates basic and advanced CAGE analyses.
• The study provides a reproducible R-code framework for CAGE data analysis.
• Guidelines are established for future CAGE studies.

Conclusions:

• The CAGEfightR package offers a robust platform for comprehensive CAGE data analysis.
• This workflow facilitates detailed investigation of gene expression regulation through TSSs and enhancers.
• The study serves as a valuable resource for researchers conducting CAGE-based studies.