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Figure 2 | BMC Molecular Biology

Figure 2

From: A novel, non-radioactive eukaryotic in vitro transcription assay for sensitive quantification of RNA polymerase II activity

Figure 2

Detection of newly synthesized RNA by primer extension and PCR. In vitro transcription was performed on the pEGFP-N1 plasmid (A) or linearized CMV-EGFP template (B). The RNA was purified, reverse transcribed and detected by PCR. A. Gel analysis of the PCR products demonstrating α-amanitin-sensitive RNA synthesis. 1- DNA ladder; 2- no α-amanitin; 3- 100 μM α-amanitin; 4- negative transcription control (without NTPs); 5- positive PCR control; 6- DNase digested DNA control; 7- water PCR control. B. The visualized PCR product amount inversely correlated with the α-amanitin concentration. 1- DNA ladder; 2- no α-amanitin; 3-5- 400, 100 and 20 μM α-amanitin, respectively; 6-8- 400, 100 and 20 μM synthetic amanitin analog 30.0346, respectively; 9- negative transcription control (without NTPs); 10- positive PCR control; 11-water PCR control.

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