A trimmed viral cap-independent translation enhancing sequence for rapid in vitro gene expression

We prepared a short (29 nucleotides) 5' UTR that enhanced cap-independent t ranslation in a wheat; germ translation system by trimming the tobacco etch virus 5' UTR. The trimmed sequence, designated as TE(37-65), was obtained from a conserved region among several potyviruses. The productivities of un capped reporter mRNAs carrying the TE(37-65) sequence were comparable to th ose of capped counterparts, in that 5-20 mu g of proteins were synthesized per 1 mL of translation reaction mixture. The ribosome that entered onto th e TE(37-65) sequence precisely initiated polypeptide synthesis at the defin ed initiation codon, which ensures rapid and efficient protein truncation a nalyses. Moreover, the TE(37-65) sequence is short enough to be involved in a PCR primer, which allows a simple method for rapid gene expression, i.e. , PCR amplification of a target gene and succeeding in vitro transcription and translation. As a demonstration, the rapid in vitro expression of rice cDNAs using the TE(37-65) sequence was also performed.