At this point, it seemed odd that only one out of the four possible combinations of Nacetyl-/N-formyl-aminoacyl transfers across either a nicked-loop or nicked-duplex was stereoselective. We synthesised 5-mer mixed anhydrides using either L-or D-AcAla as a donor strand, then added them to a nicked duplex scenario containing the same 10-mer template strand and 8-mer acceptor strand as our earlier experiments (1 eq. 5'-AcAla-pAGCGA mixed anhydride with 1 eq. 5'-UCGCUUUCCA template and 1 eq. 5'-UAAUGGAA acceptor). We identified the transfer of AcAla residues across a nicked duplex by formation of a new peak as before (starting from Ac-L-Ala 25%, starting from Ac-D-Ala 22%; Fig. ). The species responsible for this peak were collected, lyophilised and then digested with RNAse. Subsequent HPLC analysis and comparison to synthesised standards demonstrated the presence of equal amounts of L-and D-2'/3'-AcAla-ester adenosines (Fig. ).