Figure 2
In vitro editing assay of 3' UTR targets. Total hepatic RNA from Apobec-1-/- mice was incubated with increasing amounts of WT hepatic S100 extract. RNA was used for cDNA synthesis followed by PCR amplification of Apobec-1 3′ UTR targets using specific targets. (A) Endogenous Dpyd RNA editing of cytidine 119134696 was determined by poisoned primer extension. The relative mobility of the unedited (C 4696) and edited product (U 4690) is indicated to the right. Vertically is shown the sequence surrounding the editing site. The targeted cytidine is indicated in red. Upon editing, the primer extension reaction proceeds until the next C (represented in green). The 32P-labeled primer is shown in blue. (B) Endogenous Tmbim6 RNA editing of cytidine 99239051. Total hepatic RNA from Apobec-1-/- mice was incubated with recombinant Apobec-1 and ACF or with increasing amounts of hepatic WT S100 extract. C-to-U editing of cytidine 9051 was determined by poisoned primer extension. To the right is shown the sequence surrounding the editing site. The edited cytidine (9051) is shown in red. Cytidine 9043 also appears to be targeted, resulting in an extension product terminating at cytidine 9035.