I've stumbled upon this article, Novel Chemical Scaffolds for Inhibition of Rifamycin-Resistant RNA Polymerase Discovered from High-Throughput Screening, DOI: 10.1177/1087057116679994, and I don't quite understand how they used in vitro transcription assay in their study. Can someone please enlighten me? Thank you!
Purified RNA polymerase with sigma factor was combined with a special plasmid, and the 4 ribonucleoside triphosphates (NTPs: ATP, CTP, GTP, UTP). The RNA polymerase transcribed an RNA aptamer off the plasmid. The aptamer is an RNA molecule that will fold into a tertiary structure that binds the dye Malachite Green. When the dye becomes bound to the aptamer, it becomes fluorescent. With this system, they screened a compound library for RNA polymerase inhibitors. The result of an inhibitor of RNA polymerase was a reduced production of the aptamer transcript, and therefore less Malachite Green binding, and lower fluorescence.
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