Fig. 3

Quantitative analysis of RNA transcription by qRT-PCR. In vitro transcription was performed, and the resulting RNA was reverse transcribed into cDNA, followed by qRT-PCR quantification of the cDNA. a The template DNA was serially diluted 10-fold for seven times and subjected to qRT-PCR. The cycle thresholds (CTs) were plotted against the expected copy numbers of the template DNA, showing a linear relationship (Y = − 3.677x + 40.12, r² = 0.9628). This standard curve was then used for the calculation of RNA copy number generated from transcription reactions. The graph represents the combined results of three independent experiments. Error bars represent standard deviation. b PESDNA or HNDNA was titrated with C. elegans nuclear extract in the in vitro transcription system. The RNA copy numbers generated from transcription were plotted against the amounts of C. elegans nuclear extract used in the reactions, followed by fitting with the Michaelis-Menten model and the non-linear least-squares method in Excel spreadsheets. The graph represents the combined results of three independent experiments. Error bars represent standard deviation