Transcription in vitro of T3 DNA by Escherichia coli and T3 RNA polymerases. Analysis of the products in cell-free protein-synthesizing system.

Abstract

RNA isolated from Escherichia coli infected by bacteriophage T3 was sedimented through sucrose gradients. The size distributions of the mRNA species coding for three T3‐specific enzymes, the S‐adenosyl‐l‐methionine cleaving enzyme (adenosylmethioninase), phage RNA polymerase and lysozyme, were then investigated by using the fractionated RNA to program a cell‐free protein‐synthesizing system followed by enzymatic analysis of the translation products. Identical analyses of transcripts produced in vitro by E. coli and T3 RNA polymerases were performed and it was found that E. coli RNA polymerase, in conjuncton with termination factor (ϱ), was capable of synthesizing early mRNA species (mRNA for adenosylmethioninase and phage polymerase) apparently identical in size and coding capacity to those isolated from infected cells. Transcripts synthesized by the T3 RNA polymerase, while consisting predominantly of late RNA species, were found to contain functional mRNAs for adenosylmethioninase, phage RNA polymerase and lysozyme.The transcription patterns obtained are consistent with the assignment of one promotor for the region situated near the left terminus of the genome.