The evolution of RNA structural probing methods: From gels to next-generation sequencing.

RNA molecules are important players in all domains of life and the study of the relationship between their multiple flexible states and the associated biological roles has increased in recent years. For several decades, chemical and enzymatic structural probing experiments have been used to determine RNA structure. During this time, there has been a steady improvement in probing reagents and experimental methods, and today the structural biologist community has a large range of tools at its disposal to probe the secondary structure of RNAs in vitro and in cells. Early experiments used radioactive labeling and polyacrylamide gel electrophoresis as read-out methods. This was superseded by capillary electrophoresis, and more recently by next-generation sequencing. Today, powerful structural probing methods can characterize RNA structure on a genome-wide scale. In this review, we will provide an overview of RNA structural probing methodologies from a historical and technical perspective. This article is categorized under: RNA Structure and Dynamics > RNA Structure, Dynamics, and Chemistry RNA Methods > RNA Analyses in vitro and In Silico RNA Methods > RNA Analyses in Cells.