Fig. 7. In vitro gRNA-directed U-insertion and U-deletion activities of LtREL1–TAP purified material. (A) U-insertion in vitro activity of the LtREL1–TAP purified material. Fraction 8 from Figure 5 was incubated with pre-annealed RNA substrates for the pre-cleaved insertion assay in the presence of 200 µM UTP and 20 µM ATP, and products were separated on a 15% acrylamide–urea gel. The bridged RNA substrates are shown schematically above. Guiding nucleotides are indicated in bold. ‘0’ indicates no guiding nucleotide. ‘CCC’ indicates three non-guiding C nucleotides. The locations of the ligated edited products are indicated, as are the 5′ fragments with 1–3 Us added. Control, no protein added. (B) U-deletion in vitro activity. The same fraction as in (A) was incubated with RNA substrates for pre-cleaved deletion assay. The bridged substrate which should guide the deletion of two Us is shown schematically.