A trans-activating RdRp is required for cRNP activity.

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A trans-activating RdRp is required for cRNP activity. A trans-activating RdRp is required for cRNP activity. In vitro activity assays of purified NA cRNPs (c) and vRNPs (v) in the absence of rNTPs as a negative control to indicate template RNA (Template) or (A) in the presence of rNTPs but absence of primer (–Primer), presence of ApG dinucleotide primer (ApG), or capped RNA (β-globin mRNA), (B) in the presence of rNTPs and ApG dinucleotide primer without (–Pol) or with (+Pol) added recombinant influenza virus RdRp purified from insect cells. Templates and transcription products were analyzed by primer extension using primers specific for positive sense (mRNA and cRNA) or negative sense (vRNA) NA-specific viral RNAs as described for Fig 2A. c* denotes primer extension analysis in the absence of vRNA-specific primer. Small arrows indicate products of vRNP activity, and large arrows indicate products of cRNP activity that form a doublet. Note that the weak vRNA-specific signals observed in the Template and –Pol cRNP lanes represent small amounts of vRNA that copurify with cRNPs. Ashley York et al. PNAS 2013;110:45:E4238-E4245 ©2013 by National Academy of Sciences