Hybrid plasmids containing the pyruvate dehydrogenase complex genes and gene-DNA relationships in the 2 to 3 minute region of the Escherichia coli chromosome.
A sample of colonies from the Clarke-Carbon ColE1-Escherichia coli DNA plasmid gene bank was screened by conjugation for complementation of the lipoamide dehydrogenase lesion of a deletion strain lacking all components of the pyruvate dehydrogenase complex, delta (aroP aceE aceF lpd). Two ColE1-lpd+ hybrid plasmids were identified: pGS2 (ColE1-ace lpd+; 24 kb) and pGS5 (ColE1-lpd+; 14 kb). Enzymological studies confirmed that pGS2 expressed all the activities of the pyruvate dehydrogenase complex, whereas pGS5 expressed the lipoamide dehydrogenase and acetyltransferase activities (the latter from a ColE1 promoter). These and other plasmids were used to construct a 47-site (15 enzymes) restriction map for a 24.2 kb segment of bacterial DNA in the nadC-lpd region. A further 13 sites (six enzymes) were defined in a 5.4 kb sub-segment containing the lpd gene. lambda phage derivatives containing specific fragments were constructed and used in transduction studies which located the ace and lpd genes in a 7.78 kb sub-segment flanked by AccI and NruI sites.
D010582:Bacteriophage lambda / Q000235:genetics; D002874:Chromosome Mapping; D002876:Chromosomes, Bacterial; D003227:Conjugation, Genetic; D004262:DNA Restriction Enzymes; D004269:DNA, Bacterial / Q000235:genetics*; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics* / Q000648:ultrastructure; D005798:Genes, Bacterial; D005838:Genotype; D006824:Hybridization, Genetic; D010957:Plasmids; D011768:Pyruvate Dehydrogenase Complex / Q000235:genetics*; D014161:Transduction, Genetic
D004269:DNA, Bacterial; D011768:Pyruvate Dehydrogenase Complex; D004262:DNA Restriction Enzymes