uORF Identification Using Ribosome Profiling for Gene Expression Control

The patent introduces ribosome profiling data as an intermediary to bridge the gap between simple sequence scanning and accurate uORF identification. By using ribosome occupancy patterns as a mediator, the method achieves high identification accuracy without requiring complex computational models, as the experimental data directly highlights functional uORF regions.

The patent replaces traditional mechanical sequence-matching algorithms with a data-driven approach using ribosome profiling. Instead of relying on rigid rule-based systems that fail with non-canonical features, the method uses empirical ribosome occupancy data to identify uORFs, achieving higher accuracy while maintaining computational efficiency.

The patent performs preliminary identification and validation of uORFs using ribosome profiling data before applying gene editing. By pre-characterizing the uORF sequences, locations, and functional significance, the method enables precise targeting of editing tools, thereby increasing polypeptide expression while minimizing off-target effects through accurate guide RNA design.

The patent employs feedback mechanisms where ribosome profiling data is used to verify successful uORF disruption and measure the resulting change in downstream ORF expression. This feedback loop allows for optimization of editing strategies and confirmation that desired expression changes are achieved without unintended consequences.