Skip to main content
. 2006 Jun;80(11):5482–5493. doi: 10.1128/JVI.02735-05

FIG. 7.

FIG. 7.

Transactivation, but not splicing, of P41-generated transcripts was dependent on B-AAV Rep and adenovirus. (A) B-AAV P41 transcripts. 293 cells were transfected, in the presence (+) or absence (−) of Ad5 (MOI of 5), with the following plasmids, which are diagrammed in Fig. 7C: B-AAVRepCap[1/2]TR, which contains half of the right-hand ITR which includes an RBE and the TRS (lanes 1 and 4); B-AAVRep(−)Cap[1/2]TR (in which Rep ORF was terminated, lanes 2 and 5); and B-AAVRep(−)Cap[1/2]TR, together with a Rep-supplying plasmid (B-AAVRepSM) (lanes 3 and 6). Total RNA (10 μg), isolated 36 to 40 h after transfection, was protected by the B-AAV RP probe (nt 1841 to 2039), as diagrammed. (B) A-AAV P38 transcripts. 293 cells were transfected, in the presence (+) or absence (−) of Ad5 (MOI of), with the following plasmids, which are diagrammed in panel C: A-AAV[1/2]TRRepCap (lane 1,4), which contains the right half of the A-AAV left-hand end ITR and the RepCap region; A-AAV[1/2]TRRep(−)Cap (lanes 2 and 5), in which the Rep-encoding ORF was terminated, and the plasmids A-AAV[1/2]TRRep(−)Cap and A-AAVRepSM (a A-AAV Rep supplying plasmid) (lanes 3 and 5). Total RNA (10 μg), isolated 36 to 40 h after transfection, was protected by the A-AAV RP probe (nt 1690 to 1885), as diagrammed. Plasmid C1GFP (Clontech) was cotransfected as an internal control, and RNA generated from this plasmid was protected by a probe to the GFP coding region (40). The levels of P41 (or P38) RNA (Unspl+Spl) was normalized to GFP RNA, and the activation fold (Act. Fold) is presented. The ratio of spliced P41 (or P38) transcripts to unspliced transcripts (Spl/Unspl) is also shown. Quantifications of the activation fold and the ratio of Spl/Unspl RNAs are shown with the standard deviations, and the values are averages of at least two independent experiments.