In Vivo Genome-Wide RNA Structure Probing with Structure-seq

In vivo genome-wide RNA structure probing provides a global view of RNA structure as it occurs in the cell and can assist in elucidating important functional aspects of RNA structure. Structure-seq2 provides high-quality data on transcriptome-wide RNA structure in vivo but contains numerous steps that require technical precision. In this chapter we present the steps needed to produce high-quality structural data with a focus on controls and troubleshooting. Structure-seq2 can be applied to numerous organisms including plants, humans, and bacteria and is amenable to a wide variety of RNA-modifying chemicals including DMS, glyoxal, and SHAPE reagents. Notably, the data generated by the method highlighted here can be readily analyzed using our StructureFold2 computational pipeline.