FIG. 1.

CAT reporter gene-expressing constructs with structured and linear leaders. (A) Structured leaders: S-A, synthetic shunt-competent leader shown with numbering every 10 nt. Restriction sites used for further modifications are represented by lowercase italics. S and ΔS have uORF A or the 5′-proximal sequence, respectively, deleted. S-cat has an uORF extended by the insertion of 492 nt derived from the C-terminal part of CAT. Linear leaders: L-A and L, derivatives of S-A and S, respectively, with deletion of the stem sequences (nt 79 to 120). All uORFs are boxed, ATG codons are indicated in bold, and those in CAT are underlined. Dashes represent deletions, and @ indicates the stem structures. Arrows depict sequences targeted with the antisense oligonucleotides used in the primer extension assay (primers 1 to 3). (B) Schematic representation of constructs. (C) CAT expression in wheat germ in vitro translation system; resolution by SDS-PAGE (15% gel). Translation efficiency, migration of CAT, and its cat derivative are indicated. The control has no exogenous RNA added to the system.