The construction of new vehicles for the cloning of transcription termination signals.
We have constructed new plasmids that can be used to clone transcription terminator containing DNA fragments between the first gene of the tryptophan (trp) operon and the tetracycline resistance (tet) gene. Both genes are under control of the trp promotor. Therefore the presence of a transcription termination signal on cloned fragments can be monitored by a decrease in expression of the tet gene. The plasmids contain cloning sites at different distances from the translation start signal. Consequently a cloned DNA fragment can be translated in the three possible reading frames, offering the opportunity to distinguish terminators from translation polarity (pseudo-terminators). The usefulness of the plasmids was shown by the cloning of the trp terminator and of a pseudo-terminator located in the trpB gene.
D001483:Base Sequence; D003001:Cloning, Molecular; D004262:DNA Restriction Enzymes; D004274:DNA, Recombinant / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D005796:Genes; D005838:Genotype; D009154:Mutation; D009876:Operon; D010957:Plasmids; D014158:Transcription, Genetic; D014364:Tryptophan / Q000235:genetics*
D004274:DNA, Recombinant; D014364:Tryptophan; D004262:DNA Restriction Enzymes