T7 RNA polymerase transcription of Escherichia coli isoacceptors tRNA^(Leu)

The T7 promoter and the gene encoding tRNAleu were synthesized chemically and cloned into plasmid pUC19, where the gene was transcribed in vitro with T7 RNA polymerase purified. It was found that spermidine had a negative effect on T7 transcription. The amount of transcribed tRNA was about 250 times that of the template DNA under optimal reaction conditions. The unmodified tRNAleu1 and tRNAleu2 had similar leucine accepting ability, which however was only one fourth that of the native modified ones. This result indicated that the modified nucleotides of tRNAleu played some though not key roles in its aminoacylation.

The T7 promoter and the gene encoding tRNALeu were synthesized chemically and cloned into plasmid pUC19, where the gene was transcribed in vitro with T7 RNA polymerase purified. It was found that spermidine had a negative effect on T7 transcription. The amount of transcribed tRNA was about 250 times that of the template DNA under optimal reaction conditions. The unmodified tRNA1Leu and tRNA2Leu had similar leucine accepting ability, which however was only one fourth that of the native modified ones. This result indicated that the modified nucleotides of tRNALeu played some though not key roles in its aminoacylation.