Figure 8.

CdiA‐CT^STECO31 is an anticodon nuclease. A. tRNA_UUC ^Glu sequence showing the hybridized reverse transcriptase (RT) primer and oligonucleotide standards used to map the CdiA‐CT^STECO31 cleavage site. B. Primer extension analysis. RNA was isolated competition co‐cultures and cells intoxicated by intracellular CdiA‐CT^STECO31 expression. Samples from in vitro nuclease assays were also analyzed. The neutralizing effect of CdiI^STECO31 was tested where indicated. The reverse transcriptase (RT) primer was radiolabeled, hybridized to tRNA_UUC ^Glu, and extended with reverse transcriptase. Reactions and radiolabeled marker oligonucleotides were run on a 50% urea, 15% polyacrylamide gel and visualized by phosphorimaging. C. Position of CdiA‐CT^STECO31 cleavage within the tRNA_UUC ^Glu anticodon loop. Modified nucleotides are highlighted in red. [Colour figure can be viewed at http://wileyonlinelibrary.com]