RNA structure is important for RNA function and regulation, and there is growing interest in determining the RNA structure of many transcripts. Here we provide a detailed protocol for the parallel analysis of RNA structure (PARS) for probing RNA secondary structures genome-wide. In this method, enzymatic footprinting is coupled to high-throughput sequencing to provide secondary structure data for thousands of RNAs simultaneously. The entire experimental protocol takes ∼5 d to complete, and sequencing and data analysis take an additional 6–8 d. PARS was developed using the yeast genome as proof of principle, but its approach should be applicable to probing RNA structures from different transcriptomes and structural dynamics under diverse solution conditions.
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This work was supported by National Institutes of Health (R01-HG004361). Y.W. is funded by the Agency of Science, Technology and Research of Singapore. H.Y.C. is an Early Career Scientist of the Howard Hughes Medical Institute.
Y.W. and H.Y.C. are named as inventors on a patent application filed on the PARS method by Weizmann Institute and Stanford University.
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Wan, Y., Qu, K., Ouyang, Z. et al. Genome-wide mapping of RNA structure using nuclease digestion and high-throughput sequencing. Nat Protoc 8, 849–869 (2013). https://doi.org/10.1038/nprot.2013.045
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