This information can dramatically improve secondary structure prediction 6, 7, 8. For decades, chemical and enzymatic probing have been among the most common and powerful assays available to obtain structural information on RNA at nucleotide resolution 2, 3, 4, 5. Knowledge of RNA structure in vivo therefore provides important insights regarding the evolution and function of biological systems.
Numerous classes of RNA perform key biological functions via folding into diverse structures. RNA is of central importance in gene regulation, catalysis and the origin of life 1. Our universally applicable method opens the door to identifying and exploring the specific structure-function relationships of the multitude of low-abundance RNAs that prevail in living cells. We reveal the effects of proteins on 25S rRNA, 5.8S rRNA and U12 snRNA structure, illustrating the critical importance of mapping RNA structure in vivo. Interestingly, in contrast to mammalian U12 snRNAs, the loop of the SLIIb in U12 snRNA is variable among plant species, and DMS/SHAPE-LMPCR determines it to be unstructured. We probe the structure of low-abundance U12 small nuclear RNA (snRNA) in Arabidopsis thaliana and provide in vivo evidence supporting our derived phylogenetic structure. DMS/SHAPE-LMPCR achieves attomole sensitivity, a 100,000-fold improvement over conventional methods. Here we introduce DMS/SHAPE-LMPCR to query the in vivo structures of low-abundance transcripts. However, the structures of all but the few most abundant RNAs are presently unknown in vivo.
RNA structure plays important roles in diverse biological processes.
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