T7 RNA polymerase is the most widely used enzyme in RNA synthesis, and it is also used for RNA labeling in position-selective labeling of RNA (PLOR). PLOR is a liquid-solid hybrid phase method that has been developed to introduce labels to specific positions of RNA. Here, we applied PLOR as a single-round transcription method to quantify the terminated and read-through products in transcription for the first time. Various factors, including pausing strategies, Mg2+, ligand and the NTP concentration at the transcriptional termination of adenine riboswitch RNA have been characterized. This helps to understand transcription termination, which is one of the least understood processes in transcription. Additionally, our strategy can potentially be used to study the co-transcription behavior of general RNA, especially when continuous transcription is not desired.
Keywords: PLOR; riboswitch RNA; single-round transcription; transcription termination.