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The Tyrosine tRNA Promoter

Hans Küpper, Roland Contreras, H. Gobind Khorana, Arthur Landy


The DNA of the transducing bacteriophage φ80psuIII+ has been used in recent years in studies of the transcription of the tRNA1Tyr gene by the RNA polymerase in vitro (Daniel et al. 1970; Ikeda 1971; Zubay, Cheong and Gefter 1971). The transcript containing the tRNA sequence was invariably found to be much larger (up to 1000 nucleotides) than the expected tRNA molecule. On the other hand, Altman and Smith (1971) isolated a tRNA1Tyr precursor of 129 nucleotides from E. coli cells infected with φ80psuIII+. This precursor contained a 5′ pppG end group, which immediately showed that the product contained an in vivo initiation point, and second, that the initiation occurred 41 nucleotides beyond the 5′ end of the native tRNA (Altman and Smith 1971). Whether the purified RNA polymerase recognizes the same promoter in vitro and is able to initiate transcription with the same sequence as in vivo could not be clearly demonstrated in the previous work because of the background transcription of the bacteriophage genes and the length of the in vitro tRNA precursor.

A defined study free from the above complications would be possible using DNA fragments that contain only the tRNA gene with its promoter as a template for RNA polymerase. It has been shown (Landy, Foeller and Ross 1974) that the DNA of the bacteriophage φ80psuIII+ can be cut by the restriction enzymes Hind II + III in such a way that the tRNA1Tyr gene remains intact. Furthermore, upstream of the 3′ end of the tRNA...

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