Nucleotide sequence of the 5' end of araBAD operon messenger RNA in Escherichia coli B/r

Abstract

The transcription reaction in vitro provides a means of analyzing the nucleotide sequence of the mRNA of the araBAD operon. By controlling the time of synthesis, we obtained araBAD mRNA of varying lengths beginning from the 5' end. These 5' fragments were freed of lambda RNA transcripts by successive hybridizations to the sense strands of a pair of lambda ara transducing phages that carry ara genes in opposite orientations. The purified 5' fragments were ordered by their times of appearance during synchronized RNA elongation and by nearest neighbor analyses. The results, when combined with the knowledge of the NH2-terminal sequence of the product of the first cistron (L-ribulokinase gene araB), establish the nucleotide sequence of the first 69 bases at the 5' end of the araBAD operon mRNA. The AUG starter codon for L-ribulokinase is located at positions 29-31. The sequence is: 5' A-C-C-C-G-U-U-U-U-U-U-U-U-G-G-A-U-G-G-A-G-U-G-A-A-A-C-G-A-U-G-G-C-G-A-U-U-G-C-A-A-U-U-G-G-C-C-U-C-G-A-U-U-U-U-G-C-A-G-U-G-A-U-U-C-U-G-(U)-. . .3'.