Extended Data Fig. 10: Sequence alignment of DVI of the group IIB intron and the U2 snRNA of the spliceosome.

The sequence proximal to the branch-site of 129 group IIB introns were aligned using BioEdit alignment software as described in the methods. The numbering of the nucleotides follows that of the T.el4h intron. Analysis of the aligned sequences showed complete conservations of the branch-site nucleophile as an adenosine (A860). In addition, there was also complete conservation for 832 as a purine and 858 as a pyrimidine. Furthermore, the base pair formed between nucleotides 832 and 858 never deviate from Watson-Crick. The sequence alignment was used to create the consensus secondary structure and covariation matrix for the 832–858 base pair shown in Fig. 6A of the main text. This sequence conservation correlates well with the in vitro splicing assay data where only the G832A/C858U mutant maintained wild type branching activity. For the U2 snRNA alignment, sequences from 24 diverse species were selected and the portions homologous to the AGC triad and DVI of the group II intron were aligned using BioEdit alignment software. In both cases the RNA is highly conserved with only small deviations to the U2 snRNA where it pairs to the intron to form the branch helix. The alignment also shows that the homologous nucleotide to 832 of group II introns is completely conserved as an adenosine (A35) in the spliceosome. The alignment data for the U2 snRNA was used to create the consensus secondary structure and covariation matrix seen in Fig. 6A of the main text.