Figure S3. Excision of inverse toeprints from a polyacrylamide gel.

cDNA obtained from inverse toeprints was amplified by PCR after second strand synthesis and EcoRV treatment and loaded onto a 12% acrylamide TBE gel stained with SyBR Gold. The band indicated by a black triangle corresponds to inverse toeprints of ribosomes stalled at the initiation codon. Amplified double stranded cDNA corresponding to ∼90–135 bp was excised from the gel with a clean scalpel to retain inverse toeprints where ribosomes had stalled after translating 2–15 codons. cDNA was eluted from the gel by diffusion as described in the methods section.