Figure 7.

Pseudo-pseudoknots direct ribosomes to pause over the slippery site. mRNAs generated from pJD366-18 (SL) were annealed with the sequencing oligonucleotide and either J1-, R1- or no oligo (lanes 1, 2 and 3, respectively), and these were then used for in vitro translation reactions. Reactions were stopped after 10 min by the addition of cycloheximide, and reverse transcription reactions were initiated on the sequencing oligonucleotides. In parallel, control reverse transcription reactions were carried out using synthetic JD366-18 mRNA and oligonucleotides, but without in vitro translation (lanes 4–6). The positions of the slippery site, loops and stems of the pseudo-pseudoknots are indicated next to a sequencing reaction. Arrowheads indicate positions of reverse transcriptase strong stops and these are mapped to a representation of the stem–loop structure of pJD366-18.