Transposon-generated Tn10 insertion mutations at thearo genes ofEscherichia coli K-12 |
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Authors: | Antonio Cobos María F. Fernández Dr. Pablo E. Hernández Bernabé Sanz |
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Affiliation: | (1) Dpto. Higiene y Tecnología de los Alimentos, Facultad de Veterinaria, Universidad Complutense, 28040 Madrid, Spain |
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Abstract: | We have obtained a set ofEscherichia coli K-12 derivatives with transposon-generated Tn10 insertion mutations at thearo genes of their aromatic biosynthetic pathway. Bacteriophage NK561 (Tn10) has been used for transposon mutagenesis ofE. coli, strain BW545. Tetracycline (Tc)-resistant derivatives were screened by their Aro– phenotype by growth on a minimal medium with adequate requirements. Sixaro mutant types were mapped; two strains werearoA, twoaroD, onearoB oraroE, and onearoC. A selective medium and ad-cycloserine enrichment in the presence of tetracycline were used to select for Aro–, Tc-sensitive derivatives. The reversion index to aromatic-independent colonies of some derivatives was less than 2 × 10–11 per bacterium per generation. P1 transduction experiments transferred an aroA::Tn10 insertion fromE. coli BW545 to an enterotoxigenicE. coli strain from porcine origin. Derivatives of this strain beingaro, Tc-sensitive and not reverting toaro+ at a detectable frequency, and many others transduced at will, may prove their usefulness as live vaccines. |
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