The effect of alteration of 5' and 3' flanking sequences on the transcription of plant tRNA genes was analysed using an RNA polymerase III-dependent in vitro transcription system derived from nuclei of cultured tobacco cells. A TATA-like sequence and the CAA motif frequently observed upstream of plant tRNA genes, and the poly(T) stretch usually present downstream, were shown to be necessary for efficient re-initiation of transcription. The CAA motif was shown to be a transcription initiation site. Introduction of the CAA and TATA-like motifs into a gene naturally lacking them greatly enhanced transcription by promoting efficient re-initiation.
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