FIG. 4.
Toeprint analysis of a gene VII transcript. Assays were carried out as described in Materials and Methods, and samples of identical sizes from the reaction mixtures were electrophoresed on an 8% sequencing gel. In lanes containing tRNA, the full-length cDNA product reproducibly showed decreased mobility. Symbols at the top of the lanes indicate the presence (+) or absence (−) of tRNAfMet and 30S ribosomal subunits, the amounts of 30S subunits present (6.5 and 11 pmol), and incubation temperatures (37 and 42°C). Where not otherwise indicated, reaction mixtures contained 6.5 pmol of 30S subunits and were incubated at 37°C. Sequencing reactions were performed in the absence of tRNA and 30S ribosomes, with the indicated ddNTP present at 200 μM. The sequence highlights the two in-frame ATGs (boxed) and +16 positions (arrows) corresponding to each.