ANALYSIS OF THE SELENOCYSTEINE TRNA([SER]SEC) GENE-TRANSCRIPTION IN-VITRO USING XENOPUS OOCYTE EXTRACTS

An in vitro transcription system was devised using a Xenopus oocyte S1 50 extract for analyzing expression of the Sec tRNA([Ser]Sec) gene. Th e activator element, located at about -200 that is required for maxima l expression of the tRNA([Sec]Sec) gene in vivo, had no effect on tRNA ([Ser]Sec) transcription in vitro. In vitro transcription of this gene even tolerated mutations within or deletion of the PSE, showing marke d contrast with the requirements observed in vivo. However, the TATA b ox was indispensable for basal level expression of the gene both in vi vo and in vitro. The region spanning from the TATA box to the transcri ption start point (-33 to -1) was sufficient for Pol III to recognize the tRNA([Ser]Sec) gene promoter in vitro. (C) 1996 Academic Press, In c.